Restriction enzyme

Restriction enzyme digestion lab
Formal Lab Report on Restriction Digestion Analysis. Overview: In today’s lab you will use a technique called restriction enzyme mapping to determine the identity of two different plasmids (small circular pieces of DNA). ... This is similar to the use of standard curves for estimating the concentration of smaller molecules, as you learned in Lab 2 (Spectrophotometry of Proteins) of this laboratory course. There is, however, one notable difference between the standard curves of Lab 2 and the standard curve that you will construct in this exercise.
Bio 6 – Restriction Enzyme Mapping Lab
In the previous lab you learned how digest a DNA sample with a restriction enzyme and examine the resulting restriction fragments by agarose gel electrophoresis. You will use the same techniques in this laboratory for the purpose of creating a restriction map – a map of the relative positions of multiple restriction sites. The DNA molecule for which you will create the map is a circular plasmid that contains a total of 3342 base pairs (bp) of DNA
Week 1 restriction enzyme digest date needed...
The purpose of this lab is to introduce two techniques commonly used in molecular biology: restriction enzyme digestion of DNA, and characterization of DNA by. agarose gel electrophoresis. These techniques will be routinely used in other experiments throughout the rest of the semester. Restriction endonucleases are enzymes which recognize and cleave double. stranded DNA at specific sequences, and are named for the cellular strain from which they are isolated. The natural function of the enzymes is to destroy foreign DNA, while.
Bi 1x Spring 2017 | 2.2.2 Restriction enzymes
These techniques will be used over and over again in this class and certainly beyond if you choose to embark on further studies in the biological sciences. Though we are learning basic technical procedures, these exercises will still illustrate biological principles. In this introduction to lab practices, you will perform four exercises. ... Restriction enzymes are generally supplied as a given number of units. These units correspond to a metric of enzymatic activity, as specied by the manufacturer. Your digests use enzymes from NEB, which uses the following denition for a unit: “One unit is dened as the amount of enzyme...
Lab: Gel Electrophoresis and Restriction Enzymes
restriction enzymes which we will use in this laboratory are EcoRI, HindIII, and BamHI and their sequences are as follows, with the cut side indicated by the arrow. EcoRI 5’….G AATTC….3’ 3’….CTTAA G….5’. HindIII 5’….A AGCTT….3’ 3’….TTCGA A….5’. BamHI 5’….G GATCC….3’ 3’….CCTAG G….5’. ... 22. Wash hands and clean lab station. Post lab activities: Calculating the Sizes of Restriction Fragment Length Polymorphisms Mathematical formulas have been developed for describing the relationship between the molecular. weight of a DNA fragment and its mobility (i.e., how far it migrates in the gel).
Lab 5: DNA Fingerprinting | Background: Restriction enzymes
P-10 pipet, P-100 pipet, yellow micropipet tips ? colored microfuge tubes (green, blue, orange, violet, red, & yellow) ? sharpie marker to label tubes ? foam microfuge tube holder for waterbath incubation ? laboratory tape to label samples, gels. ... products of your restriction enzyme digestion. Once separated on a gel, the DNA fragments will then be visualized by DNA staining in Activity 5c. Background: Gel electrophoresis: lab technique that uses electricity and a thin gel to separate nucleic acids or proteins by size.
AP Lab Nine: Biotechnology (Restriction Enzyme Analysis of...
Restriction enzymes are enzymes that recognize specific DNA sequences in double-stranded DNA (usually a 4-6 base pair sequence) and digest DNA at these sites. The result is fragments of DNA in various, predictable lengths. ... In this lab, we will observe DNA recombination fragments of the different suspects in a crime scene using gel electrophoresis. In electrophoresis, an electrical field is used to migrate samples of the DNA strands through an agarose gel. The smaller the sequence, the faster it will maneuver through the gel.
Prelab discussion #12 | creating restriction maps
O Lab Final: Dec 7th and 8th : Bring your calculator. Will focus on assessing. your understanding of the experimental techniques covered in 421 labs. throughout the semester in terms of collecting, analyzing, and. interpreting data. ... Each plasmid contains: • Rel gene • Ampicilin resistence gene • SP6 Promoter • Origin of replication • Restriction enzyme recognition sites. Restriction endonucleases (re). Enzymes that recognize and digest DNA by cleaving phosphodiester bonds between nucleotides.
BIO 2960 Lab: Restriction Digest Computer Lab
One powerful technique in molecular biology is physically mapping DNA molecules with restriction endonucleases. In 1979, Nathans, Smith and Arber were awarded the Nobel Prize for discovering restriction enzymes and having the insight and creativity to use these enzymes to map genes. In today's lab, you will construct a very basic restriction map of the plasmid pUC19, which is a small (2686 bp) vector derived from a naturally-occurring E. coli plasmid.
9 March
Bio101- LAB 8
For this reason, restriction enzymes are valuable tools in the laboratory. Restriction enzymes are naturally occurring enzymes that cut DNA at specific sites. (Bacteria use them as defense mechanisms to destroy any invading DNA from bacterial viruses.) ... In our lab today we will simulate the process of DNA profiling using DNA digests and gel electrophoresis to distinguish multiple DNA samples, including one from a “crime scene”, and samples from four potential suspects. These techniques should allow us to determine if any of our “suspects” are a close match to the DNA left at the “crime scene”!
Restriction Enzyme Digestion
NOTES: 1= Many NEB enzymes now work in the new buffer system called CutSmart. CutSmart is basically NEB Buffer #4 and BSA combined (10X solution). Before using CutSmart, ensure your enzyme’s compatability on 2= Restriction enzyme activity is measured in “units.” One unit is defined as the amount of the enzyme required to digest 1 ug of DNA in 60 minutes. 10-fold overdigestion is recommended. In our lab, use 10 units of enzyme for DNA amounts of 1 ug or less.
Using Restriction Mapping to Teach Basic Skills in the...
This laboratory exercise has as its explicit goal the production of a restriction map of an ‘‘unknown’’ plasmid using four different restriction enzymes, though goals for student learning are much more encompassing. Restric-tion mapping is still used to compare clones [2], follow traits within and between species for ... ‘‘The restriction enzyme mapping lab was really neat and helpful in learning how to run gels and read them.’’ ‘‘I was very excited to do many of the protocols that I had only read about previously.’’ ‘‘Hands-on experience of major techniques made you feel empowered in some strange way.’’
Basic Biotechnology Kit | ENZYMES
Restriction enzymes easily and rapidly degrade unless kept frozen. Loss of activity will occur if the enzymes are subjected to warm temperatures for any length of time. When aliquoting student reagents, be careful to keep them on ice. Also, be sure to use good. sterile technique when you prepare student aliquots. ... Laboratory. Background Reading. In this lab, you will use special chemicals called restriction enzymes that act as chemical scissors to cut DNA into pieces. Restriction enzymes make two incisions, one through each of the phosphate backbones of the double helix without damaging the bases.
Case of the Crown Jewels
Data/Observation Sheets. S-8 Used in the Laboratory Activity. Restriction Enzyme Worksheet #1. S-10. ... These classroom and laboratory activities meet several of the Maryland Science Content Standards: Goal 1.0 Skills and Processes 1.3.1 The student will develop and demonstrate skills in using lab and field equipment to perform investigative techniques. * 1.2.6 The student will identify appropriate methods for conducting an investigation (independent and dependent variables, proper controls, repeat trials, appropriate sample size, etc.).
Biology 331 | Assignment - Lab Report
2) Review micropipettor technique. 3) Understand the properties of the components of a restriction digest. ... The restriction enzymes we will use in this laboratory are homodimers that recognize DNA sequences that show two-fold symmetry (Table 1). You will use linear phage lambda DNA as your substrate. The intact genome is 48,502 base pairs in length. During the lab period, you will digest this ge-nome with restriction enzymes X, Y, and Z.
Biotechnology Lab Program
Laboratory 2. pGRN + = pGRN +Hind III pGRN - = uncut pGRN (pGRN without enzyme) Include your group number and class period, on each tube, so that you can locate them for the next lab period. 3. The reaction matrix summarizes the reagents used in the restriction digest. ... INTRODUCTION. The purpose of this lab is to collect a DNA sample from the cells that line the inside of your mouth and to use this sample to explore one of the most powerful techniques in molecular biology- the Polymerase Chain Reaction, PCR. Although PCR has many applications, it is commonly used to produce many copies of a selected...
Biology 382 – Techniques in Molecular Biology – Syllabus...
isolation, followed by restriction enzyme analysis with agarose gel electrophoresis, DNA sequencing, and sequence analysis to evaluate success of the procedure. Class periods will be used flexibly: class may begin with lecture or with lab work (and short lectures interspersed during the lab; at other times we will begin with any of the following: pop quiz, demonstrations by instructor, or viewing of films or videos. ... Lab Citizenship: Safety and Courtesy Strictly following all safety rules is basic to good lab technique. ? No eating, drinking, smoking or application of cosmetics in the laboratory.
Explain the role of vectors, restriction enzymes, and DNA ligase in genetic engineering. ¦ Give examples of different types of vectors used in genetic engineering. ¦ Describe the process of transformation and carry out this procedure in the laboratory. ¦ Inoculate agar plates with bacteria. ¦ ... For example, scientists can produce large quantities of proteins that are used as enzymes in commercial manufacturing and food processing techniques. Other proteins produced by gene cloning, such as human growth hormone, are medically useful.
Summer Programs at Johns Hopkins | Pre-College Course...
Discover Hopkins: Introduction to Laboratory Research. This course will introduce students to a variety of biochemical and molecular biological laboratory techniques. These will include DNA analysis by restriction enzyme mapping, amplification of DNA segments by PCR, lipid analysis by chromatography. ... Care to contribute to an ongoing research program and start establishing your scientific legacy? Students will isolate and characterize novel bacteriophages (viruses that infect bacteria) from the environment using modern molecular biological techniques. Includes training in lab safety, and sterile technique.
14 April
Laboratory 4. diagnostic digestion of plasmid dna
Restriction enzymes, or restriction endonucleases, are proteins that recognize and bind to specific DNA sequences and cut the DNA at or near the recognition site. Restriction enzymes were originally discovered through their ability to break down, or "restrict" foreign DNA. ... Additional terms to discuss – heat inactivation, star activity, self-ligation, restriction digestion of larger DNA, incubation time, incubation temperature Exercise 3. Gel Electrophoresis (refer to Lab 1, Exercise 2). Conclusions for Laboratory 5. What did your digests tell you?
Biotechnology Explorer
This advance preparation will ensure that the labs run smoothly and that the students will understand the concepts behind each laboratory. ... The techniques introduced in this exercise form the basis of recombinant DNA technology techniques, DNA fingerprinting, and forensic DNA analysis. This kit introduces students to some important principles of genetic engineering. Specifically, the functions of restriction enzymes and their use as molecular biology tools will be stressed.
Restriction Enzyme Cleavage of DNA and Electrophoresis
PRACTICE GEL LOADING If you are unfamiliar with loading samples in agarose gels, it is recommended that you practice sample delivery techniques before conducting the experiment. 1. A small sample of gel is in the plastic dish in front of you covered with water. ... 2. Predict the number of DNA fragments and their sizes if Lambda phage DNA were incubated and cleaved simultaneously with both Hind III and Eco RI (refer to the map below). Biology of the Cell Lab (BIOL1021) Restriction Enzyme Lab.
LAB 12 | Enzyme
LABORATORY OVERVIEW In this lab, the relative location of restriction sites on a plasmid will be mapped. Each team will set up digestions and then run them on an agarose gel with standard DNA markers. The size of all resulting fragments will be estimated using the standard markers, and the plasmid map ... Construct a data table that lists the different size fragments of DNA resulting when this (these) recombinant plasmid(s) are cut with the restriction enzymes used in this lab. Once you have gel electrophoresis results, analyze them to determine which Lambda DNA fragment was inserted into the pUC18 plasmid.
Lab # 7Restriction Enzymes. General Genetics. Objectives: Introduce the students to digest genomic DNA by restriction endonucleases. Observe the results of digestion on agarose gel electrophoresis. Theoretical Basis Using Restriction Enzymes. The activity of restriction enzymes is dependent upon precise environmental conditions: PH. Temperature. Salt Concentration. Ions. An Enzymatic Unit (u) is defined as the amount of enzyme required to digest 1 ug of DNA under optimal conditions
Bio 210A | Restriction Enzyme
The system was later found to consist of enzymes that can cut foreign viral DNA and as such these enzymes would restrict the viral DNA from replicating in a bacterial cell by cutting the sugar-phosphate backbone of the viral DNA. ... Materials. Plasmid rpARA (50 ng/ ?L) Restriction enzyme mixture (BamH I + Hind III) 2.5x restriction buffer Distilled water (dH2O) P-20 micropipette and tips 1.5 ml microfuge tubes Minicentrifuge. 37°C water bath Permanent marker. Procedure. In this laboratory you will be using two restriction enzymes, Hind III and BamH I, to digest the plasmid rpARA and to cut out the 702 bp...
Digestion of DNA With | Using Restriction Enzymes
S-Adenosyl Methionine. Using Restriction Enzymes. Each restriction enzyme has its own optimal set of reaction conditions, which can be found on the information sheet provided by the supplier. ... Orange G 0.25% orange G (Sigma cat # O-1625) dissolved in 50% sucrose. Bromphenol Blue 0.25 g bromphenol blue 0.25 g xylene cyanol 1.0 ml 1M Tris, pH8 49 ml water 50 ml glycerol. Cold Spring Harbor Laboratory Restriction Digestion Buffers: From A Manual for Genetic Engineering: Advanced Bacterial Genetics.
Experiment 6 | Week 1 – Restriction enzyme digestion
In this lab we will use restriction enzymes for the second purpose. Different species of bacteria have different restriction enzymes. The enzymes are referred to by a designation of the species and strain from which they are isolated. ... Therefore, the basic idea behind this technique is quite simple. To compare two different DNA samples you digest them with the same restriction enzyme and use gel electrophoresis to examine.
Primary school programmes (pri 5 - pri 6) | DNA Lab
technique has been used to insert a gene for human insulin into bacteria for mass. production of insulin. Students will also learn the sterile lab techniques necessary to. work with bacteria, including growing bacteria colonies via plating. Practicals ... Students will understand the concept of electrophoresis and how it separates a mixture of DNA fragments. They will also learn how to predict a DNA fingerprint using restriction maps. Concepts on restriction enzyme analysis will provide the students with the knowledge of such applications.
New Page 2
Laboratory Objectives. After completion of this lab you should: be able to define and explain the terms “restriction endonuclease”, “restriction site”, “restriction digest”, "palindromic sequence“, "isoschizomer", "blunt- and sticky DNA ends". know how to set up a restriction digest to cleave DNA at defined restriction sites using defined restriction enzymes. be able to interpret the experimental outcomes of a typical restriction digest experiment using circular plasmid DNA and be able to analyze simple restriction fragment patterns after agarose gel electrophoresis.
19 March
This laboratory protocol uses the restriction enzymes BamH...
The sites labeled “BamH I” and “Hind III” represents restriction sites for these two restriction enzymes. Study the plasmid map below and locate these plasmid components. The plasmid pKAN-R carries the kanamycin resistant gene, kanr, which encodes a phosphotransferase, an en-zyme that transfers a phosphate group to the kanamycin molecule destroying its antibiotic effects. ... During this laboratory, then, you will remove the rfp gene from pKAN-R and remove the small 40bp fragment from the pARA plasmid using the same enzymes.
BIO 107 Lab # 8: Summary & Study guide
• TE buffer was used for storage. TE Buffer contains Tris/EDTA solution: Tris solution buffers the pH to protect DNA; while EDTA solution protects the DNA from degradation by DNases. EDTA removes divalent cations that are needed for enzymatic function of DNase. • This procedure is sensitive, as the DNA can be sheared ... Furthermore, DNA extraction is crucial to obtain samples for DNA fingerprinting, a technique used in forensic science, paternity testing and identification of victims. • You will extract onion DNA to freeze it for lab 9: restriction enzyme digestion of DNA followed by DNA gel electrophoresis (see p. 87).
Biology 163 laboratory. Restriction mapping of plasmid dna. (Reviewed Fall 2014). Physical mapping of genomes is an important part of modern molecular genetics. ... You will use two important molecular techniques to create a restriction map of a common cloning plasmid. In Part 1 of this investigation, you will cut the plasmid into fragments with three restriction endonucleases (a.k.a. restriction enzymes). In Part 2 you will separate the fragments according to size using gel electrophoresis.
Laboratory 1: Cloning and Construction of Recombinant DNA
The biotech revolution, however, may have been launched in 1972 by Herbert Boyer and Stanley Cohen, who developed a procedure for making “recombinant DNA”, i.e., DNA consisting of fragments from different organisms. Their technique was the basis of what is now a multibillion dollar industry. ... Cohen’s lab had developed a method of introducing plasmids containing antibiotic genes into certain bacteria, while Boyer’s group had isolated an enzyme that cut DNA in a precise, predicable, reproducible manner. These enzymes, of which hundreds have been now identified, are called “restriction enzymes”.
The ? Bacteriophage Paritcle (Virion)
Lab 4. Molecular Cloning of Viral DNA Fragments in a Bacterial Plasmid Vector. Reading: ? Appendix VIII. Restriction Enzymes ? Biology 6th ed. By Campbell et al. DNA Cloning” p. 375-383. This lab exercise generally follows the sequence of numbered steps shown in Campbell’s Fig. ... The X-gal screening technique will signal the presence of foreign DNA in the vector, but typically one wants to actually “see” the insert to absolutely validate its presence and to determine its size. This means isolating it by cutting it out of the plasmid. You will do this in the subsequent steps of this laboratory exercise.
Factors that Influence Restriction Enzyme Activity
If the sequence is known, restriction sites can be predicted with accuracy, but in the lab, an enzyme may cut more often than it should or at the wrong sites. In some cases, these unexpected results point to a problem not related to technique - for example, the sequence you have may be incorrect, or a restriction map provided by a colleague could be in error. However, there are a number of commonly-encountered situtions that influence how well restriction enzymes cut, and it is important to be aware of these for troubleshooting.
1 October
Ctgtgcaagcatgacgt gaattc gaggtcaacacatg
For this reason it is important for you to begin to gain experience with the basic fundamentals of molecular biology in the laboratory. One of the most widely publicized uses of molecular biology techniques is for the purpose of identification of criminals through what is commonly called Forensic DNA ... Each lab bench will also receive a tube containing the restriction enzymes EcoR1 and Sal1 in the appropriate buffer to allow the enzymes to function. You will pipet 5?l of the enzyme mixture into each of the tubes containing the suspects’ DNA samples and to the tube containing the crime scene sample.
Restriction enzyme analysis of dna
1. Obtain DNA already digested with the restriction enzyme. Keep on ICE. ... It is a technique for separating and analyzing mixtures of charged molecules. ... Upon completion of this lab. • Dispose of designated materials in the appropriate places. •
BIBC 103: Biochemical Techniques | Lab Schedule
Chapter 7: The New Genetics—Techniques for DNA Analysis
Before the 1980s, finding the genotype of an individual usually involved various laboratory assays for a gene product—the protein or enzyme. The cases of the ABO and Rhesus blood groups are classic examples of how one infers genotypes from the reaction of gene products with certain chemicals. ... For example, the restriction enzyme EcoRI (for E. Coli Restriction enzyme number I)7 recognizes the sequence GAATTC and slices the DNA right after the G. Restriction enzymes are used in a wide variety of techniques.
Laboratory Techniques
7 November
Restriction enzyme digests, agarose gel electrophoresis and...
All 4 of these restriction enzymes will digest at 37? C, but you will have to use specific buffers for each. If you are setting up many restriction digests with the same enzyme you can make a master mix. Practice sterile molecular biology techniques when pipetting and do not contaminate the enzyme stocks. ... You can use any restriction enzyme mapping program available on the web (see website suggestions in additional resources list on the class website). Make sure you cite in your lab write up what website you used.
Biology 475 | Biol 324 Restriction enzyme tables
To set up a restriction digestion, you must mix precise amounts of various components to get the correct amount of DNA, enzyme, buffer and other reagents. The most effective method of determining how much of what to add is to set up a restriction digest table. This handout gives you a set of hypothetical restriction digests that you should work out before coming to lab. ... - provides sodium ions and a Tris buffer which keeps the pH at the right level. 1 mg/ml BSA. - to keep the enzyme stable during the digestion. 100 mM DTT. – a reducing reagent to stabilize enzyme. 1 mg/ml DNA. - substrate for restriction enzyme.
ESC102/bme final lab report
The methods used in the experiment utilize PCR and restriction digest for medical diagnostics – this novel way of determining genetic differences between individuals is significant in its inexpensive and feasible nature. The use of PCR techniques for information-gathering and molecular analysis of clinical samples is an innovation in the ... Part 3: Restriction Digest and Electrophoresis As in Lab 3, except: Before step 1, took 15 ?L PCR product (for each of the four samples labelled "LH", "MY", "148" and "090" test tubes) and transferred 15 ?L of SacII enzyme into each, then incubated at 37oC for 1 hour.
Dna, restriction enzymes, and
Dna, restriction enzymes, and gel electrophoresis. Introduction. In this two-day lab you will explore the many properties of DNA. ... Note its appearance and consistency. 2. Place 1 ml of Tris EDTA (TE) buffer into the test tube. The Tris serves to buffer the solution at about pH 8.0 while the EDTA chelates Mg++ which is a necessary cofactor for many DNAases and thereby protects the DNA from enzymatic degradation.
Biology 6B
¦ Laboratory research — Perform routine procedures used in biological research laboratories, especially as related to the techniques of molecular biology. Demonstrate proficiency with standard protocols of lab etiquette, safety, hazardous materials handling, and documentation. ... (endo- means in). Restriction enzymes were the first DNA-altering enzymes to be isolated and used in the laboratory; in a sense, molecular biology began when a restriction enzyme was used to cut DNA in a test tube. Why do cells have.
Restriction Enzyme Analysis
Thursday, January 19 - Orientation (all students), Barnum 114. Unit I. Molecular Biology: Cloning, Transformation, Plasmid Isolation, Restriction Enzyme Analysis. ... 1/23, 1/25 Introduction to Lab Techniques; Construction of Recombinant Plasmids. 1/30, 2/1 Transformation of E. coli with Recombinant Plasmids. 2/6, 2/8. Isolation of Plasmid DNA from Transformants.
Laboratory. ... Module III report Pre-Lab: Restriction Enzymes. 4th Floor Labs. 12/13 – 12/16. HGDRP Final Quiz Discussion and Problems. Lab Report: Restriction Enzymes Lab Report: Human DNA.
6 December
Chemistry 472B | Basic Laboratory Techniques: Pipetting
Basic Laboratory Techniques: Buffers. Proteins, and especially enzymes, are generally quite sensitive to changes in the concentrations of various solution components. ... Laboratory strains typically have the restriction enzyme system inactivated to prevent the degradation of introduced plasmid DNA (for example, in K-12-derived E. coli strains, hsdRMS mutants have both restriction enzyme EcoKI and the corresponding methylase genes deleted, while hsdR17 (rK- mK+) mutants have the EcoKI gene inactivated, but retain the methylase).
Laboratory Field Trips | Clemson University, South Carolina
Mapping Plasmids Using Restriction Enzymes (3.5 hours + lunch break). Students will: • Learn the origins and uses of restriction endonucleases (enzymes). • Set up restriction digests. • Run digested DNA fragments on agarose gels. • ... Two concurrent laboratory field trips (49-64 participants) — $600. *Your DNA and Traits lab involves many techniques and reagents resulting in additional fees of 1.5x the normal rate.
18 July
Restriction fragments (pieces of DNA generated by cutting with a restriction enzyme) may be separated based on their size using a technique called electrophoresis. Electrophoresis involves applying an electrical charge to a sample. Because DNA has a negative charge, it will migrate toward the positive pole. ... Also, we want to produce a DNA fingerprint and use it to determine the identity of a person suspected of committing a robbery. Please keep in mind that a very careful and sterile technique is required to perform this lab.
Lab 7: pAMP & pKAN ligation | Cloning techniques & strategies
Objective of this lab is to create a recombinant plasmid constructed from pieces of two different plasmids joined together. ... Cloning techniques & strategies. Why use two different restriction enzymes? Using 2 enzymes to generate fragments with incompatible sticky/cohesive ends greatly improves your chances of obtaining the desired product of ligation (compared to using a single restriction enzyme).
Restriction enzymes, produced by microorganisms, are...
Molecular Biology of Life Laboratory. Restriction Enzymes. BIOL 123. ... RE vial twice. • The RE should be the last component that goes into a reaction mixture. Use of any section of this Lab Manual without the written consent of Dr. Eby Bassiri, Dept. of Biology, University of Pennsylvania is strictly prohibited.
Lab 3
Most restriction enzymes work most efficiently at specific temperatures; if it’s too cold the enzymes are not active, if it’s too hot the enzymes are destroyed. In the lab we add buffer and enzyme to PCR product, then put it in an incubator or thermocycler set at a specific temperature and let the reaction sit for several hours. ... ddH20 8.4. 8.225. FISH 543: Molecular Techniques. Table 1. Mobility range of DNA in different percentage agarose gels (taken from the Owl Easycast owner’s manual – link on webpage).
DNA Fingerprinting By Restriction Enzyme Digestion of DNA
This technique can be applied to almost any DNA sample from whole genomes to individual genes. ... Restriction Enzyme Digest • Eppendorf centrifuge tubes (sometimes called “Ep” tubes) • Racks for holding the Ep tubes • Ice buckets • Vortex mixer • Hind III (restriction enzyme) • Restriction enzyme buffer at ten fold (10X) the proper concentration needed for the reaction • 37°C water bath • Sample DNA extracted from the seized bacteriaphage particals, and DNA extracted from bacteriaphage samples taken from several labs at U.
Dna restriction analysis
and will refrigerate until next lab period when we will look at them. 9. The gels can then be placed on gel support film, which binds the gel and dehydrates it, if your instructor so chooses. INTERPRETATION: Examine your stained gel on a light box, overhead projector, or a UV box. Which restriction enzyme produced the most restriction sites on the lambda DNA? The HindIII digest of lambda DNA yields at least 6 fragments suitable for use as molecular weight standards for gel electrophoresis.
The Case of the Missing Necklace
2. The four nucleotides can create a large number of sequences. 3. DNA can be manipulated and identified by using laboratory techniques. 4. Restriction enzymes can be used to cut DNA strands. ... You will simulate these two lab processes with paper. The first process is the use of restriction enzymes. Restriction enzymes are proteins that are made naturally by bacteria. Bacteria make these enzymes to fight viruses that attack them. These enzymes cut DNA at specific places where there is a specific sequence of base pairs.
BASTYR | Biochemistry for Life Science Lab II
Lab 3. DNA Digestion by Restriction Enzyme. Lambda DNA will be digested by three different enzymes to produce DNA fragments. ... *Laboratory performance will be an evaluation of lab technique, lab preparedness, lab etiquette; which includes cleaning after oneself, keeping track of lab glassware, putting away chemicals, effort given to make for a successful experiment, and most important, following safety instructions.
Chapter 11 | Restriction Enzyme
Restriction enzymes cleave specific sites in DNA Restriction enzymes like EcoRI are frequently called 6-cutters, because they recognize a 6-nucleotide sequence. Assuming a random distribution of A, C, G and Ts in DNA, probability predicts that a recognition site for a 6-cutter should occur about once for every 4096 bp (46) in DNA. ... Handling restriction endonucleases in the laboratory The REs that we are using in the lab are highly purified (and expensive!) proteins that have been purified from recombinant bacteria.
Restriction Dig Restriction fragment Remi You have transformed bacteria with plasmid DNA You have isolated plasmid D This is the third and final section for RESULTS that will be part of your “in-lab report interpretation”. ... Determine size of fragments. Present the “map” of the plasmid in your report The steps in BLUE you will complete outsi What A restriction enzyme(s)?
Figure 8: Restriction enzymes.
There are many different types of restriction enzymes. Each type recognizes a different restriction site. In this lab, Lambda DNA, which is a commercially available DNA normally found in a virus called Phage Lambda, will be restricted with the restriction enzyme BamH1. ... Describe the major techniques used in this lab: DNA Restriction, Gel Electrophoresis, etc. Important properties of DNA directly having an impact on the extraction procedure. Clearly describe the procedural steps the way they were carried out in lab.
16 August
Cloning via Restriction Digest
2. Select restriction enzymes for your insert and vector, and determine the appropriate reaction buffers. 3. Combine the following in a microfuge tube (30 uL total volume): 2 ug DNA 1 uL Each Restriction Enzyme 3 uL 10x Buffer 3 uL 10x BSA (if recommended) x uL H2O (to bring total volume to ... Tip: Run cold water over the outside of the flask for faster cooling. Safety tip: Ethidium bromide is a known mutagen. Wear a lab coat, eye protection, and gloves when working with this chemical. In our lab we use Cybr Safe. XXXX 3. Pour the agarose/ethidium bromide solution into a casting tray with the well comb in place.
Bio 121 Lab 11
In the second part of today’s lab we will use restriction enzymes to analyze the DNA which we extracted last week. ... Each restriction enzyme recognizes and cuts a specific sequence, therefore, we can determine the number of times that the sequence recognized by a particular enzyme occurs within a DNA molecule by measuring the number of fragments which it creates (fragments created by restriction enzymes are called "restriction fragments").
3 March
R estriction e nzyme b ackground
A note on restriction enzyme names: most are named after the scientific name of the species they were isolated from; Xho I is from Xanthomonas holcicola, and EcoR I is from E. coli. Pronunciation is a little funny and sometimes varies by lab; for example, in some labs, EcoR I is pronounced "eek oh are one." In other places, you may hear "eck oh are one."
DNA Fingerprinting in the Context of Criminal Forensics
Goggles and heat-protective gloves should be used when preparing agarose gels. Lab coats should be worn at all times. Science Content for the Teacher: DNA fingerprinting is a common technique used for linking genetic material found at crime scenes with genetic material obtained from suspects. ... Preparation (Before Students Arrive): 30 minutes 1. Rehydrate DNA samples, enzymes, and mixes included in kit, according to directions listed in the kit. Day One: 45 minute class session – Restriction Digestion For the sake of simplicity in a high school laboratory environment, the samples in the included kit have...
Restriction Enzymes | ASU - Ask A Biologist
Restriction enzymes, also known as restriction endonucleases, are enzymes that cut a DNA molecule at a particular place. They are essential tools for recombinant DNA technology. The enzyme "scans" a DNA molecule, looking for a particular sequence, usually of four to six nucleotides. ... Most enzymes come in glycerol solution as a storage buffer, but enzymes don't work well in the presence of high glycerol concentration. You want to be sure to dilute the glycerol content to less than 5% to ensure proper enzymatic activity.
25 May
Forensic DNA Fingerprinting Kit
In this lab activity, students will compare band patterns produced by restriction enzyme cleavage of DNA samples when separated on an agarose gel (RFLP). The patterns in this exercise are produced from one sample that represents DNA taken at the crime scene and five samples obtained from suspects in the case. It may be important for you to point out to your students that this laboratory exercise models the more elaborate technique that is performed on complex human DNA samples.
Key Words DNA Electrophoresis Restriction enzyme...
Seating Arrangement Laboratory groups of 2 – 3 students. Maximum Number of Students 30. Key Words DNA Electrophoresis Restriction enzyme Molecular biology. 2 ... Procedures and results should be recorded in a lab notebook or written report. 7. Introduce the technique of restriction enzyme cleavage using the word analogy activity described in “How Restriction Enzyme, Probes and RFLP’s Work.”
Summer 2014 LABORATORY SCHEDULE. Title/Agenda Introduction to lab ... Lab Exam 1-Lab periods 1-14 or 1-13 (Pipetting Techniques through Enzyme Kinetics ... Issues: Genetic testing 4) Human Genetic Typing: Part C. - Restriction enzyme digests 5)...
Jordan University of Science and Technology
Electrophoresis Polymerase Chain Reaction I Polymerase Chain Reaction II Restriction Enzyme I Restriction Enzyme II Plasmid Isolation Exploring DNA Data Bases on the World Wide Web Lymphocyte culture Harvesting and Staining Karyotyping I Karyotyping II. ... Handout 9 Handout 10 Handout 11 Handout 12. Additional Notes. This Lab covers large scale of Molecular and Cytogenetic techniques in an advanced way of performing experiments.
Restriction Enzyme Digestions
Restriction Enzyme Digestions. 1. Determine which 1, 2, or 3 restriction enzymes are to be included in the digestion reaction. Check “NEB double digest finder” to ensure that all enzymes are compatible in the same buffer type. [Note: most enzymes are being shifted towards “CutSmart Buffer”]. 2. Set up reaction in an Eppendorf tube (e.g. 50 uL reaction)
BI327 - Lab Protocol | About Restriction Enzymes
©CLONTECH. BIOL 327 Molecular Biology. Lab Protocol #4. Restriction Enzyme Digestion of DNA. ... Each restriction enzyme has a set of optimal reaction conditions, which are given on the information sheet and in the catalogues supplied by the manufacturer. The major variables in the reaction are the temperature of incubation and the composition of the reaction buffer. Most companies supply 10x concentrates of these buffers with the enzymes.
1 April
MIT Lincoln Laboratory: Lab Note: Uncoiling DNA Analysis
Lab Notes. BIOCHEMISTRY Uncoiling DNA Analysis. Posted November 2010. ... The core of the technique is restriction enzymes, which are produced naturally in bacteria and protect their hosts by binding to specific known sequences on foreign DNA strands and cutting the strands into fragments. Schwoebel, a member of the Laboratory's Chemical, Biological, and Nanoscale Technologies Group, recognized that if they could get the restriction enzymes to bind but not cut the DNA and attach fluorescent markers to the enzymes, they should be able to create optically detectable patterns of binding, or...
4 December
Genomic DNA is very large.
Objectives: A) To determine the rough location of restriction sites of an unknown restriction enzyme and B) to use this information to determine the identity of this restriction enzyme. Introduction: Genomic DNA is very large. For example, the human genome contains over 1 billion ... Your instructor will give you the exact protocol in the pre lab lecture. The general procedure will be to add the appropriate buffer to the DNA sample. Different enzymes require different buffer conditions, pH, salt concentration, and the type of buffer (phosphate verses acetate for example) all are important to enzyme activity.
Experiment 2
For many DNA manipulations such as restriction enzyme analysis, subcloning and agarose gel electrophoresis, the simple methods are sufficient. The high quality preparations are required for most DNA sequencing, PCR manipulations, transformation and other techniques. ... Results and discussion section of your lab report: Plot the logarithm of the number of base pairs of the standard (?/Hind III/EcoRI DNA) on the y-axis vs. distance migrated (x-axis).
Lab Manual: Part I | Restriction enzyme and plasmid
Restriction enzyme and plasmid. Objectives: DNA technology has a daily impact to Biology as well as to our everyday life. In the previous experiment, we have tested whether drug resistance can be transferred by transformation using a plasmid containing resistance gene. In this experiment we will conduct restriction analysis of this plasmid. The objectives of this lab is to get familiar with the some basic techniques and terms frequently used for genetic engineering.
Lab 7: Molecular Biology
Currently, sophisticated techniques are being exploited for isolating DNA, cloning genes, and determining how genes function. To comprehend fully these topics, you must understand some of the basic principles of DNA analysis. In this series of labs you will learn some simple methods for isolating and analyzing DNA. You will first extract plasmid DNA from bacteria. You will then digest (break apart) samples of your DNA with restriction enzymes in preparation for the analysis of the DNA by agarose gel electrophoresis.
Restriction Enzymes
A particular restriction enzyme will typically cut an organism's DNA in to many pieces, from several thousand to more than a million! There is a great deal of variation in restriction sites even within a species. Although these variations do not have phenotypic expression beyond the base sequences themselves, the variants can be considered molecular "alleles," and they can be detected with sequencing techniques.
13 June
Bios 313 Background: Restriction Enzymes
Since restriction enzymes can require different buffer conditions, some strategy must be used to do double digests. The preferred method is to simultaneously digest with both enzymes in a compatible buffer. This method can be used even if one enzyme is not fully active (e.g., 75% active). ... "Dirty" DNA contaminated with protein, salts, and RNA is not cut by some enzymes. DNA from sources that methylate adenine or cytosine residues may not be digested (methylated sensitive, Table 6, pp. 126-134 Lab Fax or a catalogue appendix). Some strains of E. colimethylate the N(6) position of adenine in the sequence...
8 December
My Webspace files
24 June
Zool 3300
Zool 3300. Genetics. Laboratory Exercise. R. ... DNA. Before the lab: Read pages 419- 425 of Snustad & Simmons (fourth edition) for a description of restriction enzymes and their uses.
Restriction analysis of plasmid pBR322
Among the most important technical advances in molecular biology/genetics was the discovery and use of restriction endonucleases in the 1970's. These enzymes cleave double stranded DNA at specific base pair recognition sequences. ... During the next two weeks each lab group will conduct a set of restriction digests of a plasmid, electrophorese the digested DNA and appropriate molecular weight marker standards through an agarose gel, and visualize the restriction fragments in the gel via EtBr staining.
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18 March
16S Ribosomal RNA Sequencing (Theory) : Microbiology...
Agarose Gel Electrophoresis. Electrophoresis is a technique used in the laboratory for separating charged ... The DNA to be analyzed is first digested to completion with a restriction enzyme. ... Powered by Amrita Virtual Lab Collaborative Platform [ Ver 00.10. ]
23 January
Microbial Genetics 428L/528L - Laboratory Course
In addition, the accurate use of micropipettors is an often overlooked but critical skill. Most of you have performed these techniques in prior laboratory courses and are proficient. Therefore, in this exercise you have the opportunity to demonstrate your proficiency to us. In lieu of a lab report, your plates will be scored for ... See Appendix 2 at end of exercise regarding molecular weight standards. 7. MIC et al. 428/528L Experiment No. 4 APPENDIX 1: RESTRICTION ENZYMES (RESTRICTION ENDONUCLEASES). Many molecular techniques are based on the now simple notion of cutting and joining DNA molecules.
Refer to your previous guidesheet (lab #6) for the details of preparing another enzyme extract and the steps of the basic assay procedure. Before returning to the peroxidase, though, you'll see some visually dramatic examples of protein behavior in connection with some important lab techniques: a. effect of pH on proteins in solution b. effect of an organic solvent on proteins in solution c. centrifugation as a separation technique.
18 April
Laboratory | Exercise 1 - Measurements and Lab Techniques
Lake-Sumter Community College, Leesburg Laboratory Manual for BSC 1010C. 3. Exercise 1 - Measurements and Lab Techniques. Introduction In scientific experiments, observation and accurate measurements are essential. ... ---GXXXXAATTC-CTTAAXXXXG---. ---AXXXXAGCTT-TTCGAXXXXA---. A DNA Restriction Map provides the exact locations of the recognition sites for a particular restriction enzyme on a DNA sample. The map indicates distances from the origin where the enzyme cut the DNA.
Lab 4: Nucleic Acid- DNA, Restriction Digest, and Ligation
Once “unwound”, the DNA is easy to isolate from the rest of cell material using simple lab techniques. ... Most known restriction enzymes come from prokaryotic organisms. They were first isolated and purified from E.coli in the early 1970’s. These enzymes are called restriction enzymes because their DNA-cutting ability degraded foreign DNA that entered the host. They “restricted” the presence of foreign DNA in the bacteria.
Recombinant DNA Technology
Lecture/Lab Syllabus. Winter 2017. " Access-Ability Services (AAS) serves as a liaison and resource to the KCC community regarding disability issues, promotes equal access to all KCC programs and activities, and makes every reasonable effort to provide appropriate accommodations and assistance to students with disabilities. ... You should be able to lean the use of DNA as a molecular means for identification: 1. Principles of Southern hybridization and Northern hybridization. 2. Techniques rationale of restriction enzyme mapping. 3. Principles of DNA finger printing.
How Stable Are Diluted Restriction Enzymes?
Molecular biologists routinely use restriction enzymes as key reagents for a variety of applications including genomic mapping, restriction fragment length polymorphism (RFLP) analysis, DNA sequencing, and a host of recombinant DNA methodologies. Few would argue that these enzymes are not indispensable tools for the variety of techniques used in the ... Restriction enzymes of high purity are often stable for many years when stored at -20°C. In order to maximize the shelf life of less stable enzymes, many laboratories utilize insulated storage containers to mitigate the effects of freezer temperature uctua-tions.
Introductory Biology Laboratory
The lab practical part will carry approximately 1/3 of the total exam grade and will focus on important laboratory techniques. The format of the written test will be primarily short answers and may include diagrams and illustrations. ... E1: Bioinformatics No labs on Sep 7, 8, 9, 10, 11 E2: Microscopy E3: Microbial Techniques E4: Eukaryotic Cell Divisions Analysis of E3 results E5: Restriction Enzyme Digest and Plasmid Mapping.
Activity Summaries
Restriction enzyme analysis is used to determine whether students have the GGCC or GGGC sequence. Advance notice required for this lab. ... Other objectives include being able to match model Diatoms with Diatoms on fixed slides and discussing how structure and function of Diatoms benefits survival. Download Diatom Lab Summary. ELISA (1 Day, 50 minutes or longer class period, Grades 10-12): Students use ELISA techniques to diagnose disease exposure (demo only- students are not exposed to any disease).
26 May
Untitled Document
Enzyme assays. Antibody applications and Western Blotting. Protein purification. Students will learn practical, hands on methods used in the lab: Restriction digestion of DNA plasmids and plasmid mapping. Purify Genomic DNA. ... Laboratory Schedule (subject to adjustment as required). Readings are assigned from Metzenberg text. Week. Lab. Text/Problems/Data analysis. 8/24. Safety, Pipetting, Handling Enzymes, Sterile Technique, Qualitative Assays.
3 December
Genetics Laboratory – PCB 3063L
It will cover basic lab techniques learned throughout the course. The test will include physical manipulation of laboratory tools along with short answer and multiple choice questions about their use. ... - Lab Report 1 Final Draft Due - Grades: Participation 7, Quiz 6 (on lab 6), Data Analysis 6 March 23th/24th – LAB 8: Gene action: Synthesis of B-galactosidase in E. coli. - Grades: Participation 8, Quiz 7, Data Analysis 7 March 30th/ 31st – LAB 9: DNA Restriction enzyme digestion and polymerase chain reaction Part 1.
Lab 7 | Restriction analyses
Learning Objectives. To understand the process and purpose of purifying plasmid DNA using a commercial kit, To gain familiarity with restriction enzymes and how they interact with DNA, To be able to construct a restriction map and relate fragment size to plasmid template, and. ... Objective: To purify the plasmid DNA from overnight growths of colonies that resulted from the TOPO® cloning/transformation and to analyze the plasmid via restriction analysis and gel electrophoresis. Protocol. Part I. Miniprep of the Plasmid DNA Prior to the start of the lab, the TA will have pelleted ~5 mL of an overnight growth of a...
15 April
Bacterial transformation with recombinant dna
In this lab we are going to learn some basic techniques and concepts used to clone DNA molecules. A DNA molecule (or gene) is said to be cloned if it is contained in a vector DNA molecule from which the cloned DNA can be readily isolated. ... Plasmids also contain restriction enzyme sites in specific locations. These sites make it possible to open the circular plasmid and insert foreign DNA. In addition, many plasmids also have differential markers to allow for the identification of recombinant plasmids containing a DNA insert.
Rao Research Group
Protocol › General Techniques › DNA Primers. ... List of our general lab protocols. ... A restriction enzyme recognition sequence. A strong ribosome binding site (RBS) (sometimes you will use the ribosome binding site contained in your vector).
28 December
Restriction enzyme analysis
Each restriction enzyme has a set of optimal reaction conditions, which are given on the information sheet supplied by the manufacturer. The major variables are the temperature of incubation and the composition of the buffer. Although the temperature requirements are fairly strict, the differences between buffers are often only slight. ... If the restricted DNA is to be purified, extract once with phenol/ chloroform, once with chloroform, and precipitate the DNA with ethanol. Notes: Restriction enzymes are expensive!
Restriction Digestion and Analysis of Lambda DNA
Western Connecticut State University Biochemistry Lab CHE 431 Spring 2006. EXPERIMENT 10: Protocol & Analysis Page 7 of 7. restriction sites you observed of the restriction enzymes used in this experiment (PstI; EcoRI; and HindIII). ... Why or why not? o Which of the fragments below do you think you visualized? o What do you think would happen if you accidentally put two restriction enzymes into a single digest reactions? This technique is called a “Double Digest”. o Why would someone run a “Double Digest”?
Biotechnology Explorer
This advance preparation will ensure that the labs run smoothly and that the students will understand the concepts behind each laboratory. ... The techniques introduced in this exercise form the basis of recombinant DNA technology techniques, DNA fingerprinting, and forensic DNA analysis. This kit introduces students to some important principles of genetic engineering. Specifically, the functions of restriction enzymes and their use as molecular biology tools will be stressed.
Isolation and Electrophoresis of Plasmid
Prior to lab you should be able to: o Explain what “cloning” a gene accomplishes for a geneticist. o Describe what a plasmid is. o Describe the function of the three essential features of all cloning plasmids. o Explain how electrophoresis of DNA works. o Explain how electrophoresis can be used to determine the size of a fragment of DNA. ... Figure 2. Basic steps to generate a recombinant plasmid. The plasmid DNA starts off as a closed circular molecule. To insert a foreign piece of DNA, the plasmid is digested with a restriction enzyme that cuts the double stranded DNA at the “cloning site”.
Examples: Tasks: Ecology (Enzyme penny lab) (Authentic...
Enzyme Penny Lab (Formal Lab Report). 1. For the introduction section of your lab report, you will need to write a couple of sentences describing how enzymes work on substrates (you may need to consult your textbook if needed). ... Explain. 7. What happened to the enzyme's active site during trial II? How? 8. Why specifically does an enzyme not work as well if its active site is changed? 9. What environmental factors can affect enzyme shape? List 2. 10. What affect did inhibitors have upon reaction rate?
24 December
Enzyme Lab - Ex. 4
Enzyme - General Information. In laboratory exercise 4 you investigate five enzymes: catalase, amylase, lipase, pepsin, and trypsin. As an enzyme works it combines with its substrate and converts it to product(s). You will monitor the activity of the enzymes by observing changes in the amounts of substrate and products. ... Return to the beginning of this section Return to "Lab Help" Return to the Biology "Home Page".
21 February
Lab topic 4: enzymes
Procedure You will set up four sample tubes to illustrate how this reaction works and to examine the effects of enzyme concentration on the rate of activity. Remember when using a spectro-photometer, you must first zero the instrument using a blank (see Lab Topic 2). The blank consists of everything in the ... After three minutes you will measure absorption. Use 470 nm as your wavelength. (see Lab Topic 2 for review of spectrophotometer use). Record the absorbance of each sample tube in Table 4.1. Use your results to construct a graph of absorbance (enzyme activity) vs. enzyme concentration.
Observations of
§ Transform bacteria with jellyfish gene (GFP) § Study gene selection and regulation (amp/ara) § Restriction enzyme and ligation concepts § Advanced lab techniques § Possible extentions (GFP chromatography) § Complete in two 45 minute lab sessions. Pros and Cons. +Highlights the central molecular framework of biology: (DNAaRNAaProteinaTrait).
Biological Techniques: DNA extraction, restriction enzyme...
Biological Techniques: Spectrophotometry, enzymatic assay, bacterial growth. Research Mentoring Project. ... The day began with lectures from the undergraduate instructors on micropipetting, performing DNA restriction enzyme digests and agarose gel electrophoresis. Then, the high school students were placed into groups to complete the lab, which taught biology principles by working to solve a DNA murder mystery.
27 August
Does | In-Lab Writing Assignment (Week 1)
Week 2: • Pre-lab, students use data from the (Kass, et al., 2007) paper to describe the restriction fragment polymorphisms for the Alu element they have amplified. • In-class, students perform restriction enzyme digestion of their PCR products. • In-class students perform gel electrophoresis, photograph their gels, and begin to analyze results. ... However, a cheaper and just as effective way to determine which polymorphism you have uses two other techniques, restriction enzyme digestion and gel electrophoresis. These can be done right here in your laboratory.
Optimizing Restriction Site Placement for Synthetic Genomes
Many techniques for manip-ulating DNA make use of unique restriction sites [6, 7]. In particular, subcloning is an important method of inserting a new sequence between two dierent unique restriction sites. Thus a genomic sequence which contains unique restriction sites at regular intervals will be easy to manipulate in the laboratory. ... SiteFind is a tool which seeks to introduce a restriction enzyme as part of a point mutation using site-directed mutagenesis [13]. However, SiteFind considers the much more restricted problem of introducing a single restriction site into a short (< 400b) sequence...
CHM-4304L | Laboratory Procedure
To this end, the following major learning outcomes shall apply: Students are expected to understand basic techniques in biochemistry. 3. Laboratory Procedure. ... January 27 or 28, 2009. Lab 3: Digestion of plasmid DNA by restriction enzymes and DNA electrophoresis.
Restriction rxns | Restriction Enzymes come in 50% glycerol
1/10 final volume. DNA insert (often a PCR product) or plasmid. Restriction Enzyme 2.5-3 U/mg of DNA to be cut. Pipet to mix before and after enzyme is added. Incubate at temperature suited to the restriction enzyme being used (usually 37 C). Restriction Enzymes come in 50% glycerol. Make sure [glycerol] is <5% in total rxn. Efficient ligation of insert DNA into appropriately cut plasmid
21 March
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Links to the page contain: National Policy On The Transfer Of Scientific, Technical And......
15 December
Hamilton Smith 1973 Nobel Prize. Cutting DNA. • Restriction endonucleases (restriction enzymes) – sticky ends – blunt ends. • Nomenclature. – EcoRI – E = genus (Escherichia) – co = species (coli). – R = strain – I = # of enzyme. Some restriction enzymes.
Biol Chem 243 Overview
This course meets on Tuesday from 1 to 5 and on Thursday from 2- 5. The actual hours for the lecture portion of the class will be flexible within the hours listed above. Due to the nature of the molecular biology laboratory, students must be able to put in occasional hours outside of normally scheduled lab times in order to complete some of the procedures. ... restriction enzyme digestion, gel electrophoresis.
25 February
Digestion of Plasmid DNA
Each restriction enzyme requires specific reaction conditions for optimum activity. One of the most important reaction conditions which varies between different restriction enzymes is the salt (usually NaCl) concentration. Enzyme buffers are specifically formulated to provide the salt concentration for optimal enzyme activity. It is important, therefore, that the correct buffer solution is used for a particular restriction enzyme.
13 January
Title: Restriction enzyme digestion of...
Restriction enzyme(s): 10X Reaction Buffer(s)
Chapter 9 — Biotechnology and Recombinant DNA
Cannot digest the bacterial (host) DNA because of the presence of methylated cytosines. iv. Purified forms of bacterial restriction enzymes are used in genetic engineering. v. A specific restriction enzyme always recognizes and cuts DNA at a very specific nucleotide sequence of the DNA molecule. vi. ... It cannot be used to amplify an entire genome. III. Techniques of Genetic Engineering. a. Making recombinant DNA in the lab occurs outside a living cell. i. Once the recombinant DNA has been made, it must be put back into a cell. b. DNA can be inserted into a cell by: i. Transformation.
Restriction Digest - GreenWiki
We are borrowing the protocol from Knight:Restriction_Digest. Restriction Enzymes (NdeI, XhoI), from NEB. EcoR I buffer. BSA. Deionized, sterile H2O. Example - 50 ?L reaction. 100 ?L reactions are also common especially if your DNA to be cut is dilut...
22 May
Lauren Lee, Steven
Because of the known success in research on GAPDH, learning all of the lab techniques necessary for working in a biochemistry and molecular biology based lab was made easier. Figure 3: Materials and Methods. This flow chart shows a step by step explanation of how and what we did. In Restriction Digestion we specifically used Bgl II restriction enzyme. Results and Discussion. We lysed the cells, using a lysis buffer, to isolate genomic DNA from Arugula and Sage.
Restriction Map
There are hundreds of known enzymes called restriction endonucleases that cleave DNA at very specific sites. For example the enzyme BamHI recognizes the sequence GGATCC and cuts the DNA between the two G's. If just one base is changed in the sequence (say GGTTCC) then the enzyme will not cut the DNA. The 1978 Nobel Prize in Medicine was awarded for research on Restriction Enzymes. Restriction enzymes can cut the DNA such that they leave a 5' overhang (BamHI), a 3' overhand (PvuI), or a blunt end with no overhang (DpnI).
7 January
Restriction mapping
Indicate the restriction enzyme(s) you would use to cleave the vector for each cloning experiment. Indicate any other enzymes(s) you would need. Suppose that in your cloning of fragment a, you isolate several independent transformants and extract the single stranded DNA from phage isolated from them. How could you identify clones that contained opposite orientations of fragment a without sequencing the DNA or using restriction enzymes?
7 December
Dna technology and genomics
Restriction enzymes cut DNA molecules at specific locations called restriction sites. In nature, bacteria use restriction enzymes to cut foreign DNA and protect their own DNA by methylation. Restrictions enzymes recognize short DNA nucleotide sequences and cut at specific point in these sequences. ... These chromosomes behave normally in mitosis and can carry more DNA than a plasmid. Eukaryotic hosts are capable of providing the posttranslational modifications that many proteins require. Several techniques facilitate entry of foreign DNA. electroporation microscopically thin needles.
28 July
Restriction Modification System | Enzyme
Several different types of restriction enzymes have been found but the most useful ones for molecular biology and genetic engineering are the Type II restriction enzymes. These enzymes cut DNA at specific nucleotide sequences. For example, the enzyme EcoRI recognizes the sequence ... Thus, the viral DNA is restricted in the bacterial cell by the restriction enzyme, and the bacterial DNA is modified by the methylase and is provided protection from its own restriction enzyme.
7 December
Background Information | RESTRICTION ENZYMES
In DNA forensics laboratories, the two most commonly used restriction enzymes were Hae III and Hinf I, which are 4-base and 5-base cutting enzymes. ... 1. Label microtest tubes 1 through 4 for four restriction enzyme digestion reactions. Put your initials or group number on the tubes. 2. Tap all the tubes (see Quick Reference at left) on the lab bench to collect all the contents at the bottom of the tube. 3. Use an automatic micropipet to dispense 10 µl of Enzyme Reaction Buffer (Rxn Buffer) to each of four reaction tubes labeled 1 through 4.
How To Extract DNA From Anything Living
How? Cooling slows down enzymatic reactions. This protects DNA from enzymes that can destroy it. ... The DNA you have extracted is genomic, meaning that you have the entire collection of DNA from each cell. Unless you cut the DNA with restriction enzymes, it is too long and stringy to move through the pores of the gel. A scientist with a lab purified sample of genomic DNA might also try to sequence it or use it to perform a PCR reaction.
4 October
Restriction Digests
2. In the "pUC18" tube, we will digest the pUC18 plasmid with the restriction enzyme, HindIII as follows. Add: 3 microliters pUC18 DNA (1 microgram) 3 microliters 10X "M" restriction enzyme buffer 22 microliters dH2O 2 microliters HindIII enzyne 30 microliters total -> incubate for 1 hour at 37 degree. ... 7. When there is good separation of the lambda fragments, cut out the vector fragments and an insert fragment - the 4.4, 2.2, or 2.0 kb frag - with a razor blade, place frags in microcent tube, and freeze until next lab.
28 February
404 | The University of Virginia
This page does not exist! Not to worry. You can either go to The Everything Directory or try a search below.
15 December
What is a Restriction Enzyme
Objectives – Lab Materials 1. You should be familiar with conversions commonly used in molecular lab (i.e. converting between ml and ml and vice versa). 2. You should also be very familiar with the use of the micropipettor (“pipetmen”). For example you may be asked to measure 350ml with the P1000, what would you “dial-in” on the display?
Maple Syrup Urine Disease: A Study of Carrier Frequency in...
Restriction Enzyme Digest. Restriction enzymes are enzymes that cut DNA at specific points known as restriction sites. Restriction enzymes only recognize very specific base sequences to cleave in the DNA, which makes them useful when trying to cut DNA at specific locations on a chromosome. ... Further, they developed techniques for primer-specific restriction maps to locate the asparagine to tyrosine substitution at amino acid 394 (Mitsubuchi, et al., 1992).
21 December
Fatal error: 'break' not in the 'loop' or 'switch' context in...
Links to the page contain: Cytochrome P-450 Enzyme System....
14 August
Genetics - Restriction Enzyme Mapping
Subject: Restriction Enzymes. Grade Level(s): 11 12. Target Audience: AP Biology, Genetics. Materials Needed: handouts provided. Class Time: 2 days. Brief Summary: Students are given data collected from a lab. Restriction enzymes are used to cut up the DNA plasmid pBR322. After analyzing the data, students will create a plasmid that shows where each restriction enzyme cleaves the plasmid. Student Objective(s): Students will transfer data previously collected in a restriction enzyme lab to a chart.
11 January
Emboss: restriction enzymes
Search REBASE for enzyme name, references, suppliers etc. remap. Display a sequence with restriction cut sites, translation etc. restrict. Finds restriction enzyme cleavage sites. showseq.
20 December
Restriction Enzyme
Restriction Enzyme: Enzymes that recognize a specific sequence of double-stranded DNA and cut the DNA at that site. Restriction enzymes are often referred to as molecular scissors.
18 November
Biol 230 Lab Manual, Lab 8
Lab 7 will demonstrate that different bacteria, because of their unique enzymes, are capable of different biochemical reactions. It will also show the results of the activity of those enzymes. In later labs we will use a wide variety of special purpose differential media frequently used in the clinical laboratory to identify specific pathogenic and opportunistic bacteria. ... Aseptic Technique: Inoculation of broth tubes, slant tubes, and stab tubes. Return to Menu for Lab 8. A. starch hydrolysis.
24 October
Bio 216 - Course Syllabus
...intellectual and technical skills that allow you to excel in upper level lab laboratories, in summer ... 1 Mitochondrial Fractionation, Enzyme Assay 2 Protein Assay, Enzyme rate ... techniques, bacterial transformation, plasmid DNA isolation, restriction digests and PCR.
The restriction endonucleases cut the DNA backbone at specific sequences called recognition sites that are usually 4 or 6 base pairs long. Each restriction enzyme recognizes a different recognition sequence. As a result, a single piece of DNA is cut into several DNA fragments of different sizes. ... They can be stored frozen until ready for electrophoresis. . questions and conclusions. Name some measures that you took to prevent contamination of your DNA samples during this lab.
VIRTLAB: a virtual molecular biology laboratory. | Enrico Bucci
VIRTLAB contains a DNA sequence edi- to stimulate students in the biological sciences to analyse tor/analysis module that can be used by students for restric- and solve molecular biology problems using standard lab- tion enzyme analysis and DNA translation. VIRTLAB has oratory techniques (e.g. restriction enzyme digestions, been built so that each student has his/her own working analytical and preparative agarose gels, DNA cloning bench (Figure 1A) and freezer box.
8 June
Cutting and Splicing DNA Molecules | Restriction Enzymes
Restriction enzymes work by degrading foreign DNA. Restriction enzymes recognize and cut the sugar phosphate backbone of the DNA molecule at specific sites on a strand of DNA. The restriction enzyme recognition site is determined by a specific sequence of nucleotides. ... There are several techniques to insert recombinant DNA molecules into cells but the most common involve the use of a vector.
2 April
History of Restriction Enzymes Meeting
A lively and productive meeting on The History of Restriction Enzymes was held October 19-21, 2013, at Grace Auditorium, Cold Spring Harbor Laboratory (CSHL). ... A summarization of the meeting by Stu Linn, which included commentary on the importance of meetings, collaborations, and short visits to other labs; the importance of "small science"; and "some hot and future items" for restriction and modification studies.
16 November
New Technologies for | restriction enzyme
Two-dimensional electropho-resis of proteins follows from the techniques of one-dimensional electrophoresis and incorporates new methods of analyzing protein spots on the. gel. In several of the DNA methods, restriction enzymes are used to fragment genomic DNA be-fore sizing and ... The advantage of determining the complete genomic sequence would be that any nucleotide sequence that was identified in any laboratory could immediately be mapped to its chromosomal location. As nucleotide sequences for various parts of the genome were obtained in different lab-oratories, both common...
Lab 2: Enzyme Action (revised Fall 2009)
Lab 2 - Biol 211–Page 1 of 24. Lab 2: Enzyme Action (revised Fall 2009) In an enzyme catalyzed reaction, a substrate molecule first interacts with the active site of the enzyme, forming an enzyme-substrate complex (ES). The substrate is then converted into one or more products and then released from the enzyme.
Molecular Scissors: Restriction Enzymes 2009
Modified from: Burke, M. 2009. Laboratory Handbook: From Phenotypes to Genotypes. School of. Biological Sciences. ... The smaller the fragment of DNA, the further it will move. We can use this technique to separate DNA fragments created by restriction enzymes.
Cleavage by a restriction enzyme
In fact, proving that these bacterial enzymes cleave DNA at a specific sequence would be a tricky manner, as this research was conducted before the advent of the relatively simple DNA-sequencing techniques now available. Following on Messelson’s work, Smith set out to purify a second restriction enzyme, this time from H. influenzae, and to demonstrate that it does indeed cleave DNA in a sequence-specific manner.
4) and Thursday’s lab (Ex. 6) will be combined into one write-up Restriction Enzymes that recognize specific sequences of DNA and cleave the sugar backbone Originally named for the restriction of phage growth in certain bacteria Restriction Type II - Cuts at or near a short recognition sequence. Separate enzyme methylates the same recognition sequence. Palindromic vs. non-palindromic rec.sites Blunt vs. Sticky ends Type I - Cuts nonspecifically and far from reco Using Restricti Keep cold!
Polymerase Chain Reaction
? called "restriction enzymes“ because restrict host range for certain bacteriophage. ... 35. Theoretical Basis Using Restriction Enzymes. ? The activity of restriction enzymes is dependent upon precise environmental condtions: PH Temperature Salt Concentration Ions. ? An Enzymatic Unit (u) is defined as the amount of enzyme required to digest 1 ug of DNA under optimal conditions
Jello and Enzyme Lab
Gelatin and Enzyme Lab Enzymes are present in our body and many substances. Enzymes are substances that speed up chemical reactions. Each enzyme operates best at a particular pH and temperature. Substances on which enzymes act are called substrates. Many enzymes are named for their substrate they work with. For example, protease is an enzyme that helps break down proteins. In this lab, you will investigate the effectiveness of laundry detergents that may or may not contain enzymes.
M. Tech. in Biotechnology (BT)
Practical Advanced Genetic Engineering Lab Physico-chemical techniques Lab. ... Basic Tools & Techniques of Genetic Engineering Tools: Restriction endonuclease, DNA modifying enzymes, Different types of vectors for Cloning, sequencing and Expression of gene and high capacity vectors. Techniques: Restriction analysis (Agrose gel electrophoresis, PFGE), DNA, RNA and protein sequencing methods.
Restriction Digest
The purpose of this laboratory exercise is to use restriction enzymes to confirm the sequence of the plasmid (pYP) purified in the previous laboratory. Students will cut their purified plasmid with a series of restriction enzymes, alone and in combination. ... Copy the pUC19 sequence into the webcutter window marked "Paste the DNA sequence into the box below." Select the appropriate restriction enzymes. Include a condensed version of the printout with your report. A part of the web cutter output for analysis of restriction enzyme Dra I follows.
22 December
Restriction Enzyme Cleaving Explained
Restriction Enzyme Cleaving Explained. You'll need to understand the basic idea behind restriction enzymes to understand the simulation you'll be writing, modifying, and running. Restriction enzymes cut a strand of DNA at a specific location, the binding site, typically separating the DNA strand into two pieces. In the real chemical process a strand can be split into several pieces at multiple binding sites, we'll simulate this by repeatedly dividing a strand.
20 December
Science Weekend Investigations and Field Trips
Lab Techniques: lab safety, sterile techniques, Restriction enzyme digest, ligation, bacterial transformation, bacterial cultures, DNA purification, agarose gel electrophoresis, column chromatography, polyacrylamide gel electrophoresis, sequence comparisons on computer. ... Expectations for Students not receiving course credit: Sincere motivation to learn hands-on molecular biology lab techniques. Do interactive web tutorials in February to learn concepts of cloning, lab safety, lab techniques. (about 2 - 4 hours). Commit to full days for laboratory work, between 9am - 3pm.
11 December
Enzyme Kinetics Lab Protocol | Intro Biology
Enzyme Kinetics Lab Protocol. Learning goals for this week’s lab Students should: Understand that enzymes act as unchanged catalysts to speed up reactions in cells. Be able to estimate Vmax and Km from a graph of reaction rate vs. substrate concentration. Understand that rate can be saturated, and that it depends on the concentration of substrate (in the case where [S]>>[E].
26 August
Amylase: a sample enzyme
Objectives: After completion of this laboratory exercise you will be able to: 1. Explain the importance of enzymes in biology. 2. Explain the basic properties of an enzyme as a catalyst. 3. Discuss the effect of enzyme concentration and various inhibitors on the rate of an. enzymatic reaction. ... In this lab we will demonstrate the hydrolysis of starch to glucose using the enzyme amylase which is found in saliva and in secretions from the pancreas.
Restriction Enzymes
• A Restriction Enzyme (or restriction endonuclease) is an enzyme that cuts double-stranded DNA at specific recognition nucleotide sequences known as restriction sites. Such enzymes, found in bacteria and archaea, are thought to have evolved to provide a defense mechanism against invading viruses.
Restriction Enzyme Reaction Mix
Restriction Enzyme Reaction Mix. Buffer (10X) BSA If needed for restriction. enzyme(s) used Enzyme Note 1 DNA Note 2 Water Total. ... (2) DNA should approximate 600 ng, based on NEB recommendation of 1ug of DNA with 1 uL of enzyme in a 50 uL reaction.
Manipulating and sequencing DNA
• Cut a large piece of DNA with a restriction enzyme; • A large piece of DNA will likely contain multiple restriction enzyme cleavage sites; • Two pieces of DNA can be compared based on the number of fragments (and size of fragments) produced by a restriction enzyme digest. • ... • Isolate chromosomal DNA and break into fragments (using physical, chemical, or enzymatic techniques). • Insert these fragments into vectors(perhaps taking advantage of restriction-enzyme generated “sticky ends”).
Restriction enzymes cut DNA at specific sites
DNA methods summary. 1. Restriction enzymes cut at specic DNA sites. (N) 2. Vectors allow genes to be “cloned” and proteins “expressed”. (N) 3. Gel electrophoresis separates DNA on the basis of size. ... I diet on cod. 2. Restriction enzymes cut DNA at specific sites. • 3 types of ends: 5’ overhang, blunt and 3’ overhang • Cognate methyl transferases protect host genome from digestion. Restriction-modication systems degrade “foreign” DNA.
Making “sticky-ends” on your PCR product (insert-to-be) and...
Use PCR purification kit to remove first restriction enzyme's buffer. Then digest. with the second enzyme in appropriate buffer. • If your enzyme is EcoRI or BamHI, incubate 2-4 hours at 37°C per digestion. Run. PCR purification kit before you leave for the night, don't leave the DNA sitting with.
Restriction endonucleases | Restriction Enzyme Function
• Over 2,500 restriction enzymes have been found • Over 250 distinct specificities • Occasionally enzymes with novel DNA sequence. specificities are still found while most now prove to be duplicates (isoschizomers) of already discovered specificities. Restriction Enzyme Function. • It is generally believed that the biological function of restriction enzymes is to protect cells from foreign DNA. • Infecting DNA is cleaved (restricted) by the restriction enzyme(s) preventing it from successfully replicating and parasitizing the cell.
Restriction Enzyme Ana DNA Fingerprinti Pur 1. See the DNA 2. Become familiar with -Restriction e Applications o Used to identify bacteria and viruses based on the DNA finger printing of these organisms. ... Forensic medicine – electrophoresis is used in DNA finger printing. Lambda Phage infects E.coli. Its D Restrictions enzymes are molecular scissors Recognize specific sequence cut Gel is porous DNA fragments migrate through the gel Electric current push the DNA DNA is negatively charged because of th DNA Finge.
DNA Technology 3 Results | DNA Restriction
Starting on page 238 of the lab manual there is a good review about the analysis process. Remember that we are dealing with an Alu sequence that is present in an intron of a gene on chromosome 16. This means that the area in question is spliced out of the final protein product and not expressed. ... Also check the review starting on page 270 of the laboratory manual. It's important to remember that restriction enzymes perform an endonuclease reaction - they cleave the phosphodiester bond within a nucleotide chain. This occurs at specific sites in the genome that the enzyme recognizes.
26 October
Chapter 13 (Sample test questions)
achieve a specific goal. d. The usefulness of natural DNA recombination is determined by natural selection. e. Human interests determine the usefulness of lab DNA recombinations. DNA from different bacteria may be combined using all of the following EXCEPT a. transformation. b. plasmids. c. viruses. d. crossing over. e. all of the above. In biotechnology research, DNA fragments created by restriction enzyme action are separated from one another by: a. Crossing over b. Gel electrophoresis c. Centrifugation d. Filtering e. The polymerase chain reaction.
MCB3617/5621 Restriction Enzyme Problems. I will answer questions highlighted in red. 1. Which of the following cannot be a recognition site for a typical, Type II, restriction enzyme? Why? 5’ atcgta 3’ 5’ ttcgaa 3’ 5’ gggccc 3’ 5’ gagctc 3’ 5’ aattaa 3’. 2. EcoRI is a restriction enzyme made by certain strains of Escherichia coli. How come the enzyme doesn’t cut up the E. coli’s DNA? What is the name of the protective enzyme that prevents this? And how does it work?
BIOCHEMISTRY LABORATORY | Module 4: Enzyme Kinetics
In this laboratory experiment, you will determine the concentration of your cytochrome c sample through several measurement techniques. You will record absorption spectra at 280 and 416 nm and use extinction coefficients to estimate protein concentration. You will also contrast a standard curve using a Bradford assay. ... select appropriate restriction enzymes to insert our PCR product into a bacterial plasmid. To perform the process of PCR. Safety Wash your hands before leaving lab.
Restriction Digestion
Restriction enzymes are usually distributed fairly concentrated (10-15 units/microliter, where one unit is enough to cut one microgram of DNA per hour). If the restriction enzyme provided is diluted down, again, calculations must be done in order to assure adequate enzyme and/or time for the plasmids to be cut. Note also that environmental factors such as salt concentration, temperature, and glycerol concentration may play roles in restriction enzyme activity.
8 December
Restriction Enzymes, Gel
• Determining the sequence of nucleotides within a segment of DNA. • 1980 Chemistry: “and the other half jointly to:” • GILBERT, WALTER, U.S.A., Biological Laboratories, Cambridge, MA; and. • SANGER, FREDERICK, U.S.A., Great Britain, MRC Laboratory of Molecular Biology, Cambridge ... 8. The Major “Breakthroughs” of Molecular Biology. • Restriction Enzymes • Recombinant DNA. Techniques (“cloning”) • DNA Sequencing.
Recombinant DNA and Genetic Engineering
Precautionary measures are taken so that the recombinant bacteria won't survive outside the lab. Restriction Enzymes (Table 15.1, Fig 15.4) are produced by bacteria in response to bacteriophage (viral) infection. ... RFLPs, (Restriction fragment length polymorphisms): This techniques looks for differences in patterns when the DNA is cut with restriction enzymes. Since silent mutations and other subtle variations exist in DNA between individuals, each person has unique pattern of restriction sites in their DNA.
27 February
Computat | Restriction Enzyme
RFLP: Restriction Fragment Length Polymorphism. • A restriction enzyme cuts the DNA molecules at every occurrence of a particular sequence, called restriction site. • For example, HindII enzyme cuts at GTGCAC or GTTAAC. ... • The problem often might be formulated as recovering positions of points on a line when only some pairwise distances between points are known. (why?) • Many mapping techniques lead to the following problem: X is a set of points
Using a Single-Nucleotide | LAB FLOW
Lab: Set up PCR reactions. Post-lab: Amplify DNA in thermal cycler. Pre-lab: Aliquot HaeIII restriction enzyme. ... and Executive Director of the Dolan DNA Learning Center at Cold Spring Harbor Laboratory. • Resources: 13 animations on key techniques of molecular genetics and genomic biology, from the. award-winning Internet site, DNA Interactive.
Exercise 4: Introduction to Restriction Endonucleases
Many of them involve the use of restriction enzymes and agarose gel electrophoresis. In these experiments, you will simulate some of the steps involved in obtaining the type of evidence that appears in the courtroom in cases utilizing DNA profiling. In this laboratory activity, you will be investigating a murder case. ... 2. Add 5 µl of 5X loading dye to each reaction tube from Day 1. Mix well by either tapping the tube on the lab bench or by flicking the end of it with your finger tip. Do a quick spin in the microfuge to bring all the solution back to the bottom of the tube.
Chapter 13 | Restriction Enzymes
Recombinant DNA. Restriction Enzymes. Sticky Ends. Cloning Human Insulin. ... – Hasn’t worked in a lab for years. – Went to UC Berkeley to get a Ph.D. in biochemistry. ... • Molecular techniques produce medically important human proteins in bacteria.
Restriction Digest | Enzyme Name
9/30/13 TO Purpose Restriction digests are used to create specific sizes of DNA with “sticky ends” so that you can use those sticky ends to combine DNA together to make new plasmid variations. A restriction enzyme cuts at specific palindrome sites to create either 3’ or 5’ sticky ends. ... 5.) Clean-­?up your Restriction Digest using either: a. Gel Extraction (see protocol) b. PCR Cleanup (see protocol) c. OR you can heat inactivate the enzyme for overnight storage (see enzyme label for temp and time). 6.) Nanodrop your DNA and record your results in your lab notebook.
Simulation of Restriction Enzymes
Restriction Enzyme Cleaving Explained. You’ll need to understand the basic idea behind restriction enzymes to understand the simulation you’ll be writing, modifying, and running. Restriction enzymes cut a strand of DNA at a specific location, the binding site, typically separating the DNA strand into two pieces. ... In a genomic lab DNA doesn’t have a direction and in simulations it is often necessary to reverse a strand, e.g., change it from CGAT to TAGC. The SimpleStrand class uses the StringBuilder.reversemethod to reverse the simulated strand – note that this method changes the StringBuider object it’s called on...
2 May
Fact Sheet Describing Recombinant DNA and Elements Utilizing
One of the basic strategies of molecular cloning is to move desired genes from a large, complex genome to a small, simple one. The process of in vitro recombination makes it possible to cut different strands of DNA, in vitro (outside the cell), with a restriction enzyme and join the DNA molecules together via complementary base pairing. ... DNA sequencing is a lab technique used to determine the sequence of nucleotide bases in a molecule of DNA.
Enzyme Lab Procedures
ENZYME LAB - Part 1 - Temperature Test Procedure –. ... ENZYME LAB - Part 2 - Substrate Concentration Test Procedure –. Catalase Prep –. Get five sets of three test tubes each – designated “1mL,” “2mL,” “3mL,” “4mL,” and “5mL.”
6 April
How do phages block restriction enzymes? – Center for...
491 Research Projects Available in CPT Laboratories. How do phages block restriction enzymes? ... The well-studied bacteriophage P1 of E. coli has two anti-restriction proteins, DarA and DarB (defense against restriction), that act against several type I R-M systems found in E. coli strains. DarA inhibits the endonuclease activity of the restriction enzyme EcoA, whereas DarB prevents inhibits enzymes EcoB and EcoK.
28 August
Keywords: onion, DNA isolation, restriction enzymes...
Techniques of DNA isolation are documented that are not dependent on expensive equipment, but the DNA obtained is often contaminated with significant amounts of protein and is unusable for subsequent molecular biology manipulations. Thus, it was the goal of this project to develop a simple chromosomal DNA isolation procedure that yielded a ... It was our objective to develop an isolation procedure that could be performed using materials common to most science laboratories and would provide DNA of sufficient quality to be used in subsequent molecular biology applications (e.g. restriction enzyme digestion).
Pre-Lab Homework for Lab 5: Enzymes & Diffusion
After reading over the lab, answer the following questions. This sheet is due at the beginning of lab! 1. When we discuss the process that your cells use to control reactions, knowing the following terms will be helpful. They can all be found in your text! Catalyst. Enzyme: Substrate: 2. What does the suffix "-ase" probably mean when you see at the end of a word in a biology class? (You may need to read the lab to find this – it really is in there!)
Manipulation of Purified DNA
Analyzing the result of restriction endonuclease cleavage. A way of determining the number and sizes of the fragments is needed if restriction endonucleases are to be of use in gene cloning. Using gel electrophoresis Separation of molecules by gel electrophoresis: This technique uses differences in electrical charge to separate the molecules in a mixture. ... Only when a restriction map is available can the correct restriction endonucleases be selected for the particular cutting manipulation that is required. What restriction enzyme should be used to obtain gene B and D?
Restriction Enzyme EcoR1 | Walter and Eliza Hall Institute of...
Restriction Enzyme EcoR1. Various DNA molecular visualizations derived from x-ray crystallography and other data sets, and imbued with dynamic movement that suggest brownian motion. These molecular animations were created for a major trans-national production effort to raise awareness, educate and promote DNA science to the wider community, coinciding with the 50th anniversary of the discovery of the double helix. ... This animation visualises research published in Nature Medicine (Vol 15, Issue 8, 2009) by the laboratory of Jane Visvader and Geoffrey Lindeman.
11 February
New Page 3
[View Illustrations] | [Return to Listing].
Links to the page contain: Scale-up of Perforation Process from Laboratory Model to Bottom......
19 August
Open Access Library Journal
RFLP or restriction enzyme polymorphism is a technique in which the DNA is isolated, cut using restriction enzymes, size fractioned on gels. ... The samples were collected at random, irrespective of age and sex for each breed group. The DNA extraction was carried out at the Central Laboratory, Ministry of Science and Technology, Khartoum, Su-dan. The technique used was the Modified Guanidine Chloride protocol.
Lab III: Quantification of Enzyme Activity - Catalase
potato catalase with your human catalase. (1) Peel a fresh potato and cut the tissue into approximately pea-sized pieces (the smaller the pieces, the better they’ll blend, the less foam you’ll get, the more active your enzyme will be, and the faster lab will go). ... Keep an eye on that filter while you use the stopwatch to measure rise-times for the higher H2O2. concentrations. This laboratory was modified by M. Olney and M. Wilson from that published by B.A.D. Nichols and L.B. Cholewiak (1991) [Departments of Molecular Biology, Ecology and Evolutionary Biology, Princeton University]. http...
11 April
Biotechnology and Bioprocessing Laboratory for Chemical
The next phase of the laboratory emphasizes molecular biology techniques. The students purify plasmids from E. coli and analyze them using agarose gel electrophoresis and restriction enzyme analysis. ... As this was a first run, some of the labs did not work as expected, particularly the fermentation and enzyme immobilization. The students were also unhappy that some of the laboratory exercises required more than the three hours allotted and that in several cases, the students had to return either later or the next day to view the results.
Putman’s Biol 160 Lab 5 : Enzymology
LAB 5: Enzymology. Introduction. Every biochemical reaction that takes place within your body is controlled by a group of proteins called enzymes. Enzymes are biological catalysts, acting to speed up chemical reactions. The entire enzyme is called the holoenzyme. ... After mastery of this laboratory, including doing the assigned readings and required. laboratory work, the student should be able to: 1. Define and apply the terms. enzyme.
Young Nebraska Scientists
Restriction Enzyme Digestion and Analysis of Lambda DNA. Lambda DNA is the DNA of a bacteriophage (bacterial virus). Bacteriophages attack bacteria by inserting its DNA into the bacterial cell. ... Perform a digestion of lambda DNA with HIndIII, EcoRI, and PstI restriction enzymes. Load and run digested DNA through Electrophoresis. Generate a standard curve based on lambda HindIII DNA Standard. Determine length of all DNA fragments by comparing to the standard curve. Lab skills learned.
5 April
next. Restriction Enzymes belong to a class of bacterial enzymes (endonucleases) which recognize and cut specific DNA sequences. They are believed to have evolved as a defense against bacterial viruses. Their discovery explained the phenomenon of bacterial resistance to some viruses (the enzymes in those bacteria digest the viral DNA). ... This circumvents the natural immunity E. coli has to foreign DNA. The importance of restriction enzymes is underscored by the number of techniques it is essential for: Restriction maps, cloning, molecular markers, Southern blots, etc.
Restriction Digest Activity
Restriction Digest Activity. New England Biolabs has an excellent website that will allow you to look up a DNA sequence (by matching to a primer, for example), and visualize how that sequence will be cut with any restriction enzyme. The following description will lead you through the steps to perform this activity with the gonadotropin gene.
4 December
Arab Academy for Science, Technology & Maritime Transport
Arab Academy for Science Technology & Maritime Transport (AASTMT) a regional organization operated by Arab League and known for its undergraduate & graduate programs in Maritime Transportation, Engineering, Management, Computing & Informa...
7 August
Restriction Enzymes
gene which can be passed to offsrping) mouse that overexpresses heparanase only in the bones. Nucleotides recognized by NotI Nucleotides recognized by ClaI. Restriction Enzymes. ... After Restriction Enzyme Digestion. Linearized DNA. RE B.
Genetic Determination of ABO Genotype from
Keywords: biology, genetics, ABO, genotyping, restriction enzyme, blood typing, electrophoresis, polymerase chain reaction, buccal cell. 1. INTRODUCTION The human genome consists of a complete set of DNA that is found in almost every cell type present in the human body. ... Through performing this experiment, students will learn important laboratory techniques including PCR, gel electrophoresis, and centrifugation, as well as the foundations of ABO blood types, which are important in human physiology and immunology.
Subject Area(s)
The method chosen should be appropriate to the question being asked. (c) (2) Scientific processes. The student uses scientific methods and equipment during laboratory and field investigations. The student is expected to ... • The DNA (genetic code) from related organisms contain similar restriction sites. • Related DNA are cut into similar fragments by the same restrictions enzymes. Introduction / Motivation Comparison between multiple DNA samples can be performed by a simple technique called restriction fragment length polymorphism (RFLP).
ECO-RI DNA Restriction Enzyme Dimer
Beloit College > Chemistry > Protein Structures. This page created by George Lisensky, Beloit College. Last modified April 25, 2015.
21 January
Discussion, laboratory techniques, pictures, procedure development, and formatting. Alexis. ... Lab techniques, methods, procedure, pictures, final editing. pH Dependence of Enzymatic Activity,Harry Potter Edition. One day at Hogwarts School of Witchcraft and Wizardry, Ron Weasley and Hermione Granger just finished their Muggle Studies lesson. This one was different than their normal lessons because they had to perform, what the Muggles called, enzyme activity experiment that depended on the power hydrogen (pH).
29 September
Restriction Enzymes (Site-Specific Endonuclease)
Restriction Enzymes (Site-Specific Endonuclease). • Enzymes that recognize and cleave dsDNA in a highly sequence specific manner. • Generally recognize an inverted repeat sequence 4, 6, or 8 base pairs in length [occurrences average every 44(256bp), 46(4kbp), or 48 (65kbp)]. ... Restriction Enzymes (Site-Specific Endonuclease). • Generation of Restriction map. Analysis 6.2 kb linear DNA of unknown sequence. Use multiple restriction enzymes to deduce the restriction sites on the DNA fragment.
A list of some restriction enzymes that
The formation of single stranded DNA after a double strand break is made can be detected on Southern blots if the DNA from a time course is run on a denaturing gel. Single stranded DNA generally cannot be cut by restriction enzymes and hence runs as longer DNA segments on denaturing gels as more restriction sites become single stranded. A list of some restriction enzymes that are exceptions to this rule and are able to cleave single stranded DNA can be found in the New England Biolabs catalog.
Structural Factors that Influence the Inhibition of
One approach to answering this question would be to examine the binding of compound onto DNA by using the DNA thermal melting technique. This method was performed in the absence of any restriction enzyme to only examine interactions taking place between compound and DNA to obtain ?Tm values. ... [16] Linn, S.; Roberts, R. Nucleases. Cold Spring Harbor Laboratory, 1982, 1985; Chapter Type-II Restriction and Modification Enzymes, pp. 109-154. [17] Blackurn, G. M.; Gait, J. M. Nucleic Acids in Chemistry and Biology, 2nd Edition. Oxford University Press, 1996; Chapter 2, pp. 22-25.
Fundamentals of biotechniques
Molecular Biology and General Laboratory Techniques. ... 10 assignments Lab notebook Biotechnique reflection Oral presentation Data analysis Final Exam. ... (Combined lecture and laboratory) Nucleic Acid Module: PCR, restriction enzyme digestion.
BIL 333 Lecture I | Type II Restriction Endonucleases
vv Enzymes that cut DNA and RNA. – Exonucleases-Cut linear fragments at the ends. – Endonucleases-– cut either linear or circular nucleic acids at. sites internal to the molecule. Restriction Endonuclease. vv Endonucleases whose function depends on a specific DNA sequence. – Haemophilus influenzae = HindII. 5 GT(pyrimidine)(purine)AC 3 3 CA(purine)(pyrimidine)TG 3. Restriction Enzymes. vv. – Type I : nonspecific cleavage.
Additional information on restriction enzymes
Conversely, each type of phage can infect only selected species of bacteria, and hence have a restricted host range. Because of this effect, they became known as restriction endonucleases. ... Many bacteria contain several pairs of restriction enzymes (nucelases and DNA methylases) to increase their resistance to phage. The DNA methylases are of much less use in molecular biology than the endonucleases. As a result, the term restriction enzyme has come to refer only to the restriction endonuclease.
10 March
Gene tag construction (gtc) module
GTC Lab 4. Prerequisite information: Restriction enzymes; enzyme recognition sites. Students will gain: • understanding and practice applying molecular /biochemical principles underlying nucleic acid isolation • practice setting up basic molecular enzymatic reactions. Time: Approximately 4 hrs. ... Gtc lab 7 – instructor’s guide. Prerequisite information: Some prior discussion about Southern blotting technique. Students will gain: Practice applying Southern blotting technique. Time: Approximately 4 hours.
Restriction enzyme digestion at OHSU? See all 1 options
Enzymatic cleavage of DNA at specific sequences resulting in restriction fragments with restriction enzymes, which are enzymes isolated from bacteria that recognize specific restriction sequences in DNA. Restriction enzymes play a very important role in the construction of recombinant DNA molecules, as is done in gene cloning experiments. Filter by location: All.
22 March
Biology 3A Laboratory: Enzyme Function
Biology 3A Laboratory: Enzyme Function. Objectives • To be able to list the general characteristics of enzymes. • To study the effects of enzymes on the rate of chemical reactions. • To demonstrate the effect of some environmental conditions on enzymatic reactions such as. ... Optimum conditions may vary with different enzymes and with the location of the reaction in the body. Many enzymes also require the presence of inorganic or organic. Biology 3A Lab Enzymes.
Restriction Enzymes for AFLP’s – The Dyer Laboratory
The Dyer Laboratory. Goings on in population genetics. ... The key to the AFLP protocol is to be able to digest two restriction enzymes (RE’s) simultaneously and be able to ligate onto these sticky ends primers of known concentration. When you purchase new primers, you need to aliquot out usable volumes because repeated freeze/thaw cycles reduce RE efficiency.
27 February
Enzymes lab
Getting Started—How to Use the Online Enzyme Lab. This lab has three parts so plan your time accordingly! (Note: the lab is weighted as two lab grades because it is so long). ... On the Virtual Laboratory window (probably behind the notebook window at this point), select what type of graph you would like to plot (see second picture on the right). Select the plot type you feel is most appropriate for the data you collected (by the way, [V] represents how fast the enzyme worked in your experiment, and stands for velocity).
Enzyme Activity of Peroxidase
In lab this week we will investigate the activity of an enzyme called peroxidase. Each experiment will be conducted by a group of four students. ... During this lab exercise you will investigate the naturally occurring enzyme peroxidase. Peroxidase is a heme-containing enzyme found in peroxisomes (cellular organelles) and can be obtained from a variety of plant tissues. Each cell uses oxygen in its metabolism.
Undergraduate | BMS110 Lab (Honors Lab)
Restriction Enzyme Digestion Design Lab (1 week) • Plasmid Purification & Restriction Enzyme Digest Lab (2. ... Other topics include: biosynthesis and catabolism of biological macromolecules and related topics in biotechnology, biological nanotechnology and molecular medicine. Laboratory emphasizes hands-on experience with current techniques in biomolecular science.
Oakland Journal - Research - Restriction Enzymes
[Intro] [Tools of Research] [Mechanism of Promotor Escape] [Research Lab] [Web Links]. Restriction Enzyme Can Be Used to Limit the Length of the DNA Template Available for the Transcribing Polymerase. The DNA region that is directly contacted by the transcription complex is larger than the area required for mRNA synthesis alone. How much DNA is required for transcription can be assessed by a number of methods, one of which is limiting the length of DNA by using restriction enzymes.
10 September
Phagehunting Program
DNA Restriction Enzyme Analysis and Electrophoresis. You have a dialyzed cesium chloride banded phage stock from which you have isolated DNA by phenol-chloroform extraction and ethanol precipitation. CONGRATULATIONS!!!! Now you want to check the DNA for quantity, quality, a first estimate of ... Frame the gel so that the gel fills the screen. Push the trans-luminator button on and adjust the intensity if needed. 18. Video print two copies of your gel (one for your notebook and one for the digest book) and save the file on the Hatfull lab folder in a subfolder with your name. Be sure to label the file accurately.
Procedure altered in the following way: Peas were crushed ins DNA Detec The DNA Detectives Lab uses restriction enzymes to isolate specific fragments of DNA. The exact number and size of fragments produced by a specific restriction enzyme vary from person to person. Restriction enzymes are used to “cut” DNA molecules internally (endo) or on the ends (exo).
Lab 13: DNA Fingerprinting Using Gel Electrophoresis
KEY TERMS: DNA profile analysis/DNA profiling/DNA fingerprinting restriction cutting site/enzyme recognition sequence/restriction cleavage site agarose gel electrophoresis restriction enzyme. ... You may be asked to complete another step of the gel electrophoresis procedure listed on the bottom of C39. III. WORKSHEET: -COMPLETE AND HAND IN YOUR WORKSHEET -You can work with other students in lab to complete the worksheet.
Restriction Enzyme Digest
Additionally, restriction enzyme analysis of the PCR amplicon allowed genus-specific identification of Aspiculuris tetraptera and Syphacia spp. pinworms. Together, these newly developed assays may prove to be valuable antemortem tests for detection of pinworm infections in mice and rats, negating the need to ... Acknowledgements: This project was funded by GlaxoSmithKline, the American Society of Laboratory Animal Practitioners, and the Research Animal Diagnostic Laboratory at The University of Missouri - Columbia. PP = Pure Plasmid Prep. 5.3 MWM PP x 108 107 106 105 104 103 102 101 100.
Unit 6: Review | PH and enzyme activity continued
Unit 6: Review Enzymes Lab. Continue. Chemical Equation. Catechol oxidase reaction Read information in your lab manual. catechol + oxygen SUBSTRATE substrate. ... I. Preparation of standard tubes. Potato extract contains the enzyme, catechol oxidase. This is prepared by peeling and dicing a small potato then blending it with water. The 1% catechol is prepared in lab from a stock bottle. Continue. Preparation of control and blank tubes continued. Enzymes
Once it finds this DNA sequence it will cleave the DNA and leave it as a jagged edge. In this animation it shows the insertion of another sequence of DNA cut by the same restrictive enzyme. DNA ligase, the green complex, goes and reinitiates the phosphodieter bond between the nucleotides. Here is a link to another audio animation explaining the function of restriction enzymes: Restiction Enzymes. Problems with Restriction Enzyme. Site-selective DNA scission has become a necessary technique in manipulating genomic DNA.
24 November
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3 March
Union County College
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21 May
Student | Laboratory Procedure
These molecular scissors or “cutting” enzymes are restriction endonucleases. [Can you figure out why they are called restriction endonucleases?] Two common restriction endonucleases are EcoRI and PstI which will be provided to you in this lab procedure. ... Lesson 2 Restriction Digestion of DNA Samples. Laboratory Procedure. Upon careful observation, it is apparent that the only difference between the DNA of dif-ferent individuals is the linear sequence of their base pairs. In the lab, your team will be given 6 DNA samples.
Some Restriction Enzymes and Cleavage Sequences
Conventional, Non-molecular Techniques. Laboratory Tools. Conventional Gel Electrophoresis. Nucleic Acid (DNA-, RNA-) Based Methods. Some Restriction Enzymes and Cleavage Sequences. ... ? Restriction analysis: restriction enzymes (REs) cut DNA at sequence-specific sites, e.g.,— ? Due to genetic drift or mutations, different isolates will have variations in. the number and spacing of sites for an RE. This is called restriction-fragment length polymorphism (RFLP).
Computational Molecular Biology: Restriction Mapping
The plasmid was then partially digested with restriction enzyme X. From the partial digest, fragments of the following sizes were found: 190, 350, 540, 680, 880, 1080, 1220, 1760, 2600, 3280, 3470, 3820 & 4360 base pairs. Construct a restriction map of pBR322 showing the positions of the restriction sites for EcoRI and enzyme X. Is this a unique map?
21 February
Spe I Restriction Enzyme Displays Greater Discriminatory...
SUMMARY: Restriction enzymes SpeI and XbaI were used in a pulsed-field gel electrophoresis (PFGE) study for molecular characterization of 146 clinical Burkholderia pseudomallei isolates. The PFGE parameters were optimized to enable comparable, reproducible, and robust results. ... Two DNA plugs (2 mm each) were digested separately overnight with SpeI and XbaI (New England BioLabs, Beverly, Mass., USA). The DNA fragments were separated by electrophoresis (CHEF DR II; Bio-Rad Laboratories, Hercules, Calif., USA) in a 1z (w/v) agarose gel (Sigma, Poole, UK) in 0.5? TBE buffer for...
Enzymes and protein folding pre-lab objectives
These objectives should be completed and entered into your laboratory notebook before coming to lab. This information will be helpful for any lab quizzes/exams and is expected to be in your notebooks during notebook checks. Introduction 1. Define the following terms. Be sure you understand their role in enzymatic reactions: Enzymes catalysts Substrate active site enzyme-substrate complex activators cofactors coenzymes inhibitors. 2. List the characteristics of enzymes. 3. What is the energy of activation (EA)? How do enzymes alter EA to catalyze reactions?
BIO107: Introduction to Biotechnology
Randomizing the list will optimize your studying. Restriction Enzymes. araC Blunt ends Endonuclease Enzyme. Green fluorescence protein Nitrogenous base Nuclease Palindrome.
11 August
you obtained? Further experimentation, such as polymerase chain reaction, restriction enzyme. mapping, try to create an onion or cyanobacterial cell "from scratch", create a. DNA library, etc. ... noting current events that are related. CONCLUSION: At the completion of this lab, the students should feel more confident about. their knowledge of cellular characteristics, chemical effects on and methods to eliminate. cell parts to liberate nuclear contents, and laboratory technique and manipulation of lab tools.
6 October
Restriction Digestion Protocol ­ Introduction
Function II: Digestion. The restriction enzyme hydrolyzes (adds a hydro= water) one of the phosphodiester backbone bonds – so the chain is broken. The digestion can be either asymmetrical (shown above) or symmetrical. ... One Unit of enzyme will digest 1 microgram of DNA in 1 hour if the reaction is performed at 37C in a volume of 50ul. The Hind III restriction enzyme has the following buffer components: 1X Buffer. 10mM Tris?HCl 50mM NaCl 10mM MgCl2 1mM Dithiothreitol pH 7.9 Most active at 37C.
Biotechnology (BIOT) < Johnson County Community College
3 hrs lecture, 4 hrs lab /wk. ... Perform basic laboratory techniques related to immunology as well as explain the concepts underlying these techniques. ... Protein Methods A. Extract and purify a restriction enzyme from E. coli by ion exchange chromatography.
18 April
Separation of Mixtures Using Different... : Amrita Online Lab
Used in diagnostic laboratories for blood and urine tests. Used in dairies and home to separate butter from cream. Used in washing machines to squeeze water from wet clothes. ... Learning outcomes. Student understands the following terms: solvent extraction, chromatography, RF , centrifugation, simple distillation, fractional distillation, etc. Student acquires skills to perform experiments using the following techniques in the chemistry lab
30 September
Enzymes Lab
The purpose of this lab is to investigate the activity of the enzyme amylase within human saliva. The enzyme amylase catalyzes the breakdown of starch into glucose and maltose. This breakdown may be monitored by determining the decrease in starch quantities by employing the iodine test, or by determining the increase in reducing sugars employing benedicts reagent.
All Associates will receive a stipend increase after one year of tenure. Additional Information About National Institute of Standards and Technology (NIST). Prior affiliation includes direct full-time employment relationships with the laboratory. Research contracts with universities that provide support for graduate students or faculty who perform research on campus are not ordinarily considered to be disqualifying.
28 January
In today’s experiment you will be using a solution of lactase to test the chemical and physiological properties of this particular enzyme. Furthermore, based on the results you obtain throughout the lab, you will attempt to determine if the lactase used was extracted from human cells or bacterial cells. ... Metals can accelerate the formation of undesired disulfide bonds and can act as cofactors for proteases (enzymes responsible for hydrolyzing proteins). Chelating agents are often added to laboratory solutions to bind and remove metal ions from solution in order to slow undesired enzymatic reactions.
pGlo Restriction Map
There are usually multiple known enzyme restriction sites on either side of the uncharacterized DNA. Enzymes are biological catalysts. Catalyst denotes a substance that has the ability to increase the rate of a chemical reaction, and is not changed or destroyed by the chemical reaction that it accelerates. ... The map, sequence (and other technical information) can be found at
28 April
EXPERIMENT SIX | Restriction Mapping
The major goals for today’s lab are to (1) build an accurate consensus sequence using a technique called multiple sequence alignment; (2) use the sequence to search for our gene in a huge database of sequences held by the National Institutes of Health (NIH) using BLAST (Basic Local Alignment Search Tool). ... (3 pts) 7. What enzyme are you going to use to differentiate between the two possible orientations? Explain the logic behind using this restriction enzyme (as opposed to other enzymes, like EcoRI, etc) to determine the orientation and the expected results.
After digestion by restriction enzyme Figure. Shorter Longer f. ... DNA fingerprinting is a set of laboratory procedures That determines with near certainty whether two sam Investigator at one of the crime scen DNA Finge 1st-The DNA molecule is cut with restriction enzymes 2nd- we have to separate the fragments This is done by a technique called gel electrophoresis The DNA is placed on a tray filled with gel through.
Cystic Fibrosis Case Study: Genetic Diagnosis of Disease by...
November 22, 2013 TA: Dr. Alain Silk Lab Partners: Erika Silverman, Duaa ... The Bam HI restriction enzyme searches for the ?F508 mutation. The patient’s DNA ... Introduction: The object is to become more familiar with techniques that are often used in the field of.
Gene Mapping Techniques
The bacterial endonucleases cut the viral DNA thus restricting its further proliferation. A particular restriction endonuclease recognises a specific nucleotide sequences in DNA and cleave it. For example the restriction enzyme Hind III recognises the following DNA sequence and cuts it open as shown ... This technique provides the most direct method of visualising genes on chromosomes. A DNA probe for a particular gene is a complementary sequence for part of the gene. This is usually labelled with a fluorescent dye.
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13 December
Biotechnology Co-Listed Courses
Three hours lecture, three hours laboratory each week. Upon completion of this course, the students should be able to: • Complete calculations that are routinely encountered in a protein lab such as buffer preparation, dilutions, protein concentrations and standard curves, enzyme activity, and specific activities • Complete laboratory techniques that are commonly used in ... Demonstrate laboratory skills commonly used in molecular biology laboratories such as making solutions, DNA and RNA purification and quantization, agarose gel electrophoresis, use of restrictions and other enzymes, and PCR.
13 December
Study Guide Quiz 7
Study Guide Quiz 7. 1. Quiz 7 covers mainly the lab exercise 8 and recaps on DNA and Protein Synthesis. 2. Learn the names of techniques used to extract DNA from Wheat Germ and DNA Gel Electrophoresis to separate the variable Restriction Fragment Lengths. 3. You used detergent and meat tenderizer to break cell walls and cell membranes to release DNA. ... 6. Restriction enzymes cut DNA into Restricted fragment lengths. 7. These fragments run through agar gel at different speeds due to their different sizes and quantity.
Mathews/van Holde/Ahern 3rd Edition
Each system consists of two distinct enzyme activities: a DNA methylase and an endonuclease that catalyzes a double-strand DNA break. The type II sequence-specific endonucleases are the most widely used ones in molecular biology. Properties of each of the three types of restriction systems are discussed in the links below and are summarized in Table 25.1 ... See also: DNA Methylation, Restriction-Modification. INTERNET LINK: REBASE, The Restriction Enzyme Database.
11 March
Labnodes - The Vanderbilt Research Network
Labnodes is a system for organizing people, labs and their science. ... Bader Lab. Metabolic Physiology Shared Resource. Hormone Assay and Analytical Services Core.
15 August
BIO SCI 1993 Introduction to Biological Design and Innovation...
1. Emergency room design 2. Human Physiology 3. Medical Innovation. a. Design process b. Marketing analysis 4. Environmental Health a. Clean water considerations b. Toxicology 5. Public Health Issue a. Epidemiology considerations b. Risk factors and biostatistics 6. Molecular Biology a. Plasmid preparation b. Restriction enzymes; ligation; selection. Laboratory Techniques Literature searches and critiques Gantt charts Biostatistics Prototype of novel medical product E coli assay in water samples PCR and gel electrophoresis to identify strains of bacteria Chemical assay of water contaminants...
Fralin Life Science Institute | Optional pre-lab
It is believed that the native function of restriction enzymes was to digest foreign. DNA, in other words, to protect the microorganism from invading viral DNA. They are now used in a variety of ways in the lab. ... Technical review materials dna restriction analysis and gel electrophoresis. Adrienne Warren Chesapeake Center for Science and Technology. 1 What type of DNA are we using circular plasmid DNA.
Bioinformatics Exercises
Sequencing a genome is quite an accomplishment in itself, but it is really only the beginning of the study of an organism. Further study can be done both at the wet lab bench and on the computer. In this problem, you will use a computer to help you identify an open reading frame, determine the protein that it will express, and find the bacterial source for that protein. ... d. Some restriction enzymes generate “blunt ends,” and some generate “sticky ends.” Explain the meaning of those terms and provide an example of each. e. Go to the RESTRICT site at the Pasteur Institute (http...
A Molecular Key for the Identification
Restriction enzyme profiling, with several single enzyme digestions for a spe-cies, could allow unambiguous identification of a panel of species quickly and efficiently. PCR–RFLP–based molecular identification keys are flexible; they can be easily tailored to a laboratory, depending on the availability of restriction enzymes, allowing molecular identification of species in laboratories that do not have access to DNA sequencing. Additionally, PCR–RFLP is a fairly routine technique; the technical expertise and equipment will already be in place in most forensic laboratories.
Biology 120 Mock Lab Exam I
12. A. In lab you did an experiment with an enzyme. What is the main function of enzymes in living organisms? B. Give one other characteristic/feature of an enzyme. 13. In our ENZYME LAB: A. What was the name of the enzyme? ... 22. A. B. The plant shown here belongs to which plant division? Give one reason why this plant is restricted to moist environments. 23. A. B. The brown dots on the underside of this leaflet are called _____.
Air Force ROTC | Catalog
Credits: 1.0; May be repeated; Graded Pass/Fail Only Lec-Rec-Lab: (0-0-2) Semesters Offered: On Demand Restrictions: Permission of instructor required. AF 0230 - Precision Drill Team Techniques and skills involved in precision drill movements, including marching, rifle spinning, ceremonial sabre handling, and color guard performance. ... BL 4820 - Biochemical Laboratory Techniques I Laboratory techniques basic to biochemistry and molecular biology including protein and phospholipid determinations, purification of natural and recombinant enzymes, enzyme kinetics, polyacrylamide gel electrophoresis...
Dmitry Lyumkis Lab - Salk Institute for Biological Studies
Lyumkis Lab - Artwork. Salk researchers captured the structure of a protein complex called an intasome (center) that lets viruses similar to HIV establish permanent infection in their hosts. ... Cryo-electron microscopy, in combination with other biophysical techniques, was used to characterize the macromolecular composition, structural details, and allosteric requirements of the SgrAI restriction endonuclease. The results showed, in response to activation brought upon by invading phage, the SgrAI restriction enzyme forms run-on oligomers that are allosterically modulated by viral DNA.
2 October
Beginning Molecular Biology Laboratory Manual
Restriction Enzyme Digestion of DNA. Materials: 10X restriction enzyme buffer (see manufacturer's recommendation). ... 2 ?l of appropriate 10X restriction enzyme buffer. 0.1 to 5 ?g DNA. sterile water to a final volume of 19 ?l (Note: These volumes are for analytical digests only. Larger volumes may be necessary for preparative digests or for chromosomal DNA digests. Add 1 to 2 ?l (3 to 20 units) enzyme and mix gently.
3 August
Linkers are small, synthetic (made in the lab, or ordered from a company) DNA fragments which contain the recognition sequence for one or more restriction enzymes. After ligating on linkers, the DNA is cut with the appropriate restriction enzyme to produce ends for cloning. Random Primed DNA Synthesis for Making a “Probe” One common technique for making a probe to detect a specific DNA sequence is called “random primed DNA synthesis”.
Studying and Manipulating | restriction enzyme (cut)
A A restriction enzyme recognizes a specific base sequence in DNA (red boxes). For this and many other enzymes, the sequence is the same in the 5 to 3 direction on both strands. B Researchers use restriction enzymes to cut DNA from different sources into fragments. ... • A laboratory process by which deliberate changes are introduced into an individual’s genome. § The most common genetically modified organisms are bacteria and yeast. • Used in research, medicine, and industry • Example: To produce human insulin.
TITLE OF LAB: An Enzyme Catalyzed Reaction
OVERVIEW OF LAB DESCRIPTION: In this laboratory, we will observe the effect of an enzyme, catalase, on cellular hydrogen peroxide. ... PROBLEM What effect will catalase, a naturally occurring enzyme in the body, have on 3% hydrogen peroxide (a toxic by-product in cellular metabolism)? EXPERIMENT AND TECHNICAL OPERATION OF EQUIPMENT Materials: 3% hydrogen peroxide concentrated catalase 25 ml graduated cylinders laptop computer with Science Workshop temperature probe Two droppers (pipettes) 1 50 ml beaker.
Biotechnology Courses - Ivy Tech Community College...
Lab safety, documentation, units of measurement, instrumentation lab solutions, aseptic technique, establishing a pure culture, plasmid DNA isolation, restriction enzyme digestion, DNA agarose gel electrophoresis, PCR, DNA ligation and transformation of bacterial cells will be introduced during the course. ... These methods will include: ELISA and immunoaffinity techniques; methods for determining enzymatic activity; spectrophotometric methods; chromatographic methods; electrophoresis; light and electron microscopy. When feasible, techniques will be practiced in the laboratory setting.
15 April
One.IU | All IU Campuses
Search, Click, Done! Bringing an app store experience to IU services...
17 November
Restriction Endonucleases
Because restriction endonucleases have a conserved catalytic core, they probably all follow the same mechanism, with a few exceptions. Some restriction enzymes such as BfiI don't have a metal ion cofacter, and this will change the process. The enzyme makes one cut on each strand of the DNA. If it doesn't cut right in the middle of the restriction site ("blunt-cutter"), then it leaves single stranded DNA overhangs called sticky ends.
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