Basic PCR Technique. Presented by : Noorul Hidayah Badri
|
|
- Dwain Ball
- 5 years ago
- Views:
Transcription
1 Basic PCR Technique Presented by : Noorul Hidayah Badri
2 What is PCR? PCR is aninvitro technique which allow the amplification of a specific DNA region. PCR is like selecting a specific page from book and photocopying it millions of times. PCR is one of the most useful techniques in laboratories due to its speed and sensitivity.
3 PCR allows exponential amplification of DNA sequence - each PCR cycle theoretically doubles the amount of DNA - during PCR, an existing DNA molecule is used as a template to synthesize a new DNA strand. - through repeated cycles of DNA amplification, large quantities of DNA are produced.
4 Figure 1: PCR cycle
5 Buffer MgCl 2 Primer PCR components dntp DNA DNA Polymerase Enhancer
6 How does PCR amplify DNA? Three step: Denaturation separate the strands of the double stranded cellular DNA Annealing Lower the temperature to allow primer bind to their complementary sequences Extension polymerase synthesis DNA strands using the 3 end of the primer as the start point and the cellular DNA strand as template.
7 Parameter for optimization of PCR Many parameters need to be optimized to increase the yield and sensitivity of detection or amplification: These parameter include: - PCR thermal profile (primer annealing, extension, denaturation, number of cycles ) - MgCl 2 concentration - Primer design and concentration -Template quality and quantity
8 PCR thermal profile optimal annealing temperature depends on the length and composition of nucleotides PCR primers. Number of cycle usually between cycle number efficiency of the amplification decreased as primer & nucleotides are consumed and polymerase start to loose it activity. no increase in the desired product
9 MgCl 2 concentration Generally insufficient MgCl 2 leads to low yield and excessive MgCl 2 will result in the accumulation of nonspecific products. Optimal MgCl 2 concentration depend on the primers, DNA polymerase, template, and dntps concentration.
10 Primer design and concentration The optimum length of a primer is generally from 20 to 30 nucleotides with a melting temperature between 55 C and 65 C. If possible design the forward and reverse primer with same melting temperatures and GC content. Sequences possessing significant secondary structure should be avoided.
11 DNA quality and quantity DNAshouldbeintactandfreefrominhibitorthatcan inhibit PCR amplification. - inhibitor can be from original DNA sources ( heme from blood, melanin from hair) - inhibitor can be introduced during DNA extraction process ( phenol, sodium dodecyl sulfate (SDS), ethanol and others.) More template is not necessary better - too much DNA can cause nonspecific amplification. - too little DNA will result in little or no amplification.
12 PCR animation
13 References C.R. Newton & A. Graham (1997) PCR. BIOS Scientific Publishers. Daniel J. F. &AndersenW. R. (1999) Genetic. Wadsworth Publishing Company
14 Thank you
15 Buffer Chemical solution that provides the optimal environmental condition. 10X buffer mm KCl mmtris HCl - 15mM MgCl 2
16 MgCl 2 a necessarycofactor for DNA polymerase activity Cofactor - An organic molecule or ion (usually a metal ion) that is required by an enzyme for its activity.
17 Primer a strand of nucleic acid that serves as a starting point for DNA amplification.
18 Deoxynucleoside Triphosphate (dntps) the building blocks from which the DNA polymerase synthesizes a new DNA strand. - deoxy adenosine triphosphate (datp) - deoxy cytidine triphosphate (dctp) - deoxy guanosine triphosphate (dgtp) - deoxy thymidine triphosphate (dttp)
19 DNA Serve as template for replication
20 DNA polymerase Catalyze the synthesis of long polynucleotide chain from monomer deoxynucleoside triphosphate using one of the original parental strands as a template for the synthesis of new complementary strands.
21 Enhancers/Additives A variety of PCR additives and enhancing agents used to increase the yield, specificity and consistency of PCR reactions.
22 Table 1: Enhancer of PCR Substance Formamide 5% dimethyl sulfoxide (DMSO) < 10% Concentration Tetramethylammonium chloride (TMAC) µm Polyethylene glycol 600 (PEG) 5-15% Glycerol 10-15% Tween % Gene 32 protein 1 nm 7 deaza-dgtp Replace 75 % of dgtp with deaze-dgtp
Optimizing a Conventional Polymerase Chain Reaction (PCR) and Primer Design
Optimizing a Conventional Polymerase Chain Reaction (PCR) and Primer Design The Polymerase Chain Reaction (PCR) is a powerful technique used for the amplification of a specific segment of a nucleic acid
More informationDNA Replication. DNA Replication. Meselson & Stahl Experiment. Contents
DNA Replication Contents 1 DNA Replication 1.1 Meselson & Stahl Experiment 1.2 Replication Machinery 2 Polymerase Chain Reaction (PCR) 3 External Resources: DNA Replication Meselson & Stahl Experiment
More informationPolymerase Chain Reaction
Polymerase Chain Reaction = multiple rounds of in vitro DNA replication = a region of DNA lying between two regions of known sequence is amplified hundreds of millions of time within a matter of several
More informationRecombinant DNA Technology
History of recombinant DNA technology Recombinant DNA Technology (DNA cloning) Majid Mojarrad Recombinant DNA technology is one of the recent advances in biotechnology, which was developed by two scientists
More informationSession 3 Cloning Overview & Polymerase Chain Reaction
Session 3 Cloning Overview & Polymerase Chain Reaction Learning Objective: In this lab exercise, you will become familiar with the steps of a polymerase chain reaction, the required reagents for a successful
More informationPolymerase Chain Reaction PCR
Polymerase Chain Reaction PCR What is PCR? An in vitro process that detects, identifies, and copies (amplifies) a specific piece of DNA in a biological sample. Discovered by Dr. Kary Mullis in 1983. A
More informationPolymerase Chain Reaction (PCR)
Laboratory for Environmental Pathogens Research Department of Environmental Sciences University of Toledo Polymerase Chain Reaction (PCR) Background information The polymerase chain reaction (PCR) is an
More informationPolymerase Chain Reaction-361 BCH
Polymerase Chain Reaction-361 BCH 1-Polymerase Chain Reaction Nucleic acid amplification is an important process in biotechnology and molecular biology and has been widely used in research, medicine, agriculture
More informationCat. # R006A. For Research Use. TaKaRa Z-Taq DNA Polymerase. Product Manual. v201411da
Cat. # R006A For Research Use TaKaRa Z-Taq DNA Polymerase Product Manual Table of Contents I. Description... 3 II. Components... 3 III. Specifications... 3 IV. Optimization of Reaction Conditions... 4
More informationAh, Lou! There really are differences between us!
Name Per Ah, Lou! There really are differences between us! Introduction The human genome (the total sum of our genetic makeup) is made up of approximately 6 billion base pairs distributed on 46 chromosomes.
More informationExperiment (5): Polymerase Chain Reaction (PCR)
BCH361 [Practical] Experiment (5): Polymerase Chain Reaction (PCR) Aim: Amplification of a specific region on DNA. Primer design. Determine the parameters that may affect he specificity, fidelity and efficiency
More informationBiotechnology. Explorer Program. Serious About Science Education 5/17/09 1
Biotechnology Explorer Program Serious About Science Education 5/17/09 1 Chromosome 8: PCR TM PCR Workshop Kirk Brown,, Tracy High School; Tracy, Ca Stan Hitomi,, Monte Vista High School; Danville, CA
More informationPCR OPTIMIZATION AND TROUBLESHOOTING
PCR OPTIMIZATION AND TROUBLESHOOTING Amplification of each DNA fragment can occur only under the defined conditions which are provided by a reaction mixture. If no positive PCR result can be obtained,
More informationFactors affecting PCR
Lec. 11 Dr. Ahmed K. Ali Factors affecting PCR The sequences of the primers are critical to the success of the experiment, as are the precise temperatures used in the heating and cooling stages of the
More informationGuidelines for Preventing Contamination of PCR Reference Guidelines for Primer Design Estimation of Primer Melting Temperature
Guidelines for Preventing Contamination of PCR During PCR more than 10 million copies of a template DNA are generated. Therefore, care must be taken to avoid contamination with other templates and amplicons
More informationNEW PARADIGM of BIOTECHNOLOGY - GENET BIO. GeNet Bio Global Gene Network
NEW PARADIGM of BIOTECHNOLOGY - GENET BIO GeNet Bio Global Gene Network GENET BIO DNA AMPLIFICATION PRODUCTS GUIDE Keynote of Products Prime TaqTM DNA Polymerase Prime TaqTM Premix ExPrime TaqTM DNA Polymerase
More informationIntroduction to PCR. ABCF Training Module. Leah Kago Research Associate BecA-ILRI Hub
Introduction to PCR ABCF Training Module Leah Kago Research Associate BecA-ILRI Hub What is PCR? Polymerase Chain Reaction An in vitro process that detects, identifies, and copies (amplifies) a specific
More informationThe Polymerase Chain Reaction. Chapter 6: Background
The Polymerase Chain Reaction Chapter 6: Background Invention of PCR Kary Mullis Mile marker 46.58 in April of 1983 Pulled off the road and outlined a way to conduct DNA replication in a tube Worked for
More informationRecitation CHAPTER 9 DNA Technologies
Recitation CHAPTER 9 DNA Technologies DNA Cloning: General Scheme A cloning vector and eukaryotic chromosomes are separately cleaved with the same restriction endonuclease. (A single chromosome is shown
More informationPolymerase Chain Reaction (PCR)
Polymerase Chain Reaction (PCR) PCR protocols Polymerase Chain Reaction (PCR) A technique for the in vitro amplification of specific DNA sequences by the simultaneous primer extension of complementary
More informationPCR in the Classroom. UC Davis - PCR Workshop Friday, September 26, 2003
PCR in the Classroom UC Davis - PCR Workshop Friday, September 26, 2003 A little history In 1983, Kary B. Mullis conceived the procedure. He went on to Cetus Corp in Emeryville, CA where it was developed
More informationAmplification Products for PCR and RT-PCR
Selection guide Polymerase Hot start Comment UptiTherm DNA pol. no Most economic. Lower error rate than Taq polymerase Available in several formats, master mix including or not dntp, Mg 2+..., in gel format
More information10x Hot-Taq Buffer 1 ml 1 ml x 2ea 1 ml x 10ea. dntp Mix (each 10 mm) None / 0.2 ml None / 0.4 ml None / 0.4 ml x 5ea
HelixAmp TM Hot-Taq Polymerase (Ver. 2.0) CERTIFICATE OF ANALYSIS (1603-V01R03) Contents HelixAmp TM Hot-Taq Polymerase (Ver. 2.0) Cat. No. HT250/HT250N HT500/HT500N HT2500/HT2500N Hot-Taq (2.5 units/μl)
More informationTaq Polymerase. Data sheet. Order No.: BS (For research and in vitro applications only) Batch No.: Best before: Bio&SELL
Data sheet Order No.: BS 91.711.0100 Order No.: BS 91.711.0500 100 units 500 units (For research and in vitro applications only) Batch No.: Best before: Appearance: clear liquid Colour: transparent 1 Description
More informationPCR KIT/REAGENTS/BUFFERS/PRIMERS
PCR KIT/REAGENTS/BUFFERS/PRIMERS 114330 DNA Amplification Kit DNA amplification kit is suitable for amplification of DNA size about 100bp to 5kb. It can be also used to RAPD PCR. This kit contains all
More informationFlag illusion: stare at a fixed point on the screen for ~30 seconds. Then look at a white wall or white piece of paper. What do you see?
Midterm 2 review Flag illusion: stare at a fixed point on the screen for ~30 seconds. Then look at a white wall or white piece of paper. What do you see? RBFD: color opponency, after affects and sensory
More informationBIOLOGY - CLUTCH CH.20 - BIOTECHNOLOGY.
!! www.clutchprep.com CONCEPT: DNA CLONING DNA cloning is a technique that inserts a foreign gene into a living host to replicate the gene and produce gene products. Transformation the process by which
More informationPolymerase Chain Reaction (PCR) and Its Applications
Polymerase Chain Reaction (PCR) and Its Applications What is PCR? PCR is an exponentially progressing synthesis of the defined target DNA sequences in vitro. It was invented in 1983 by Dr. Kary Mullis,
More informationAdditional Activity: Sanger Dideoxy Sequencing: A Simulation Activity
Student Worksheet Additional Activity: Sanger Dideoxy Sequencing: A Simulation Activity LSM 6.3-7 In 1977, Frederick Sanger developed a method by which the nucleotide sequence of a DNA fragment could be
More informationAMPLIFICATION BY PCR. Target. 1. Denature. 2. Anneal primers. 3. Extend primers. Two copies of target. 1. Denature
AMPLIFICATION BY PCR Target 5 3 2. Anneal primers 3 5 1. Denature Two copies of target 3. Extend primers 1. Denature 2. Anneal primers 3. Extend primers Four copies of target PCR: First 4 Cycles PCR: Completed
More informationINTRODUCTION TO REVERSE TRANSCRIPTION PCR (RT-PCR) ABCF 2016 BecA-ILRI Hub, Nairobi 21 st September 2016 Roger Pelle Principal Scientist
INTRODUCTION TO REVERSE TRANSCRIPTION PCR (RT-PCR) ABCF 2016 BecA-ILRI Hub, Nairobi 21 st September 2016 Roger Pelle Principal Scientist Objective of PCR To provide a solution to one of the most pressing
More informationGenetics and Genomics in Medicine Chapter 3. Questions & Answers
Genetics and Genomics in Medicine Chapter 3 Multiple Choice Questions Questions & Answers Question 3.1 Which of the following statements, if any, is false? a) Amplifying DNA means making many identical
More informationPCR Amplifies Targeted Sequence
PCR Amplifies Targeted Sequence Target Sequence Supercoiled DNA Strand DNA Strand Double Helix DNA Strand Chromosome P C R PCR PCR = Polymerase Chain Reaction. A primer directed-extension reaction for
More informationTaKaRa PCR Amplification Kit
Cat. # R011 For Research Use TaKaRa PCR Amplification Kit Product Manual Table of Contents I. Description... 3 II. Components... 3 III. Storage... 4 IV. Materials Required but not Provided... 4 V. Principle...
More informationInstructions for Use Life Science Kits & Assays
Instructions for Use Life Science Kits & Assays Product specifications 1 Product specifications The innutaq HOT-A DNA Polymerase provides improved specificity and sensitivity when amplifying low-copy-number
More informationOverview. Background ~30 min. Lab activity ~50 min. DNA profiling Polymerase Chain Reaction (PCR) Gel Electrophoresis PCR
Overview Day 1: Tuesday Introduction to DNA profiling How do we use DNA to solve crimes? Background Polymerase Chain Reaction (PCR) Gel Electrophoresis Set up PCR Day 2: Wednesday Make and Run Agarose
More informationGenomic Sequencing. Genomic Sequencing. Maj Gen (R) Suhaib Ahmed, HI (M)
Maj Gen (R) Suhaib Ahmed, HI (M) The process of determining the sequence of an unknown DNA is called sequencing. There are many approaches for DNA sequencing. In the last couple of decades automated Sanger
More informationDNA: Structure & Replication
DNA Form & Function DNA: Structure & Replication Understanding DNA replication and the resulting transmission of genetic information from cell to cell, and generation to generation lays the groundwork
More informationTECHNICAL BULLETIN. SYBR Green JumpStart Taq ReadyMix without MgCl 2. Catalog Number S5193 Storage Temperature 20 C
SYBR Green JumpStart Taq ReadyMix without MgCl 2 Catalog Number S5193 Storage Temperature 20 C TECHNICAL BULLETIN Product Description SYBR Green JumpStart Taq ReadyMix without MgCl 2 combines JumpStart
More informationPolymerase chain reaction
Core course BMS361N Genetic Engineering Polymerase chain reaction Prof. Narkunaraja Shanmugam Dept. Of Biomedical Science School of Basic Medical Sciences Bharathidasan University The polymerase chain
More informationInstructions for Use Life Science Kits & Assays
Instructions for Use Life Science Kits & Assays Product specifications 1 Product specifications The innumix Standard PCR MasterMix contains all reagents required for routine high throughput PCR amplifications
More informationPCR. CSIBD Molecular Genetics Course July 12, 2011 Michael Choi, M.D.
PCR CSIBD Molecular Genetics Course July 12, 2011 Michael Choi, M.D. General Outline of the Lecture I. Background II. Basic Principles III. Detection and Analysis of PCR Products IV. Common Applications
More informationJumpStart REDAccuTaq LA DNA Polymerase. Catalog Number D1313 Storage Temperature 20 C. Product Description
JumpStart REDAccuTaq LA DNA Polymerase Catalog Number D1313 Storage Temperature 20 C Product Description JumpStart REDAccuTaq LA DNA Polymerase contains AccuTaq LA DNA polymerase, 1,2 an inert red dye,
More informationTechnical Review. Real time PCR
Technical Review Real time PCR Normal PCR: Analyze with agarose gel Normal PCR vs Real time PCR Real-time PCR, also known as quantitative PCR (qpcr) or kinetic PCR Key feature: Used to amplify and simultaneously
More informationMethods of Biomaterials Testing Lesson 3-5. Biochemical Methods - Molecular Biology -
Methods of Biomaterials Testing Lesson 3-5 Biochemical Methods - Molecular Biology - Chromosomes in the Cell Nucleus DNA in the Chromosome Deoxyribonucleic Acid (DNA) DNA has double-helix structure The
More informationhttp://fire.biol.wwu.edu/trent/trent/direct_detection_of_genotype.html 1 Like most other model organism Arabidopsis thaliana has a sequenced genome? What do we mean by sequenced genome? What sort of info
More informationThe Techniques of Molecular Biology: Forensic DNA Fingerprinting
Revised Fall 2016 The Techniques of Molecular Biology: Forensic DNA Fingerprinting **Lab coat, eye goggles and gloves (nitrile or latex) are required for this lab. You will not be allowed to participate
More informationPRODUCT INFORMATION Long PCR Enzyme Mix #K0182 500 u Lot Exp. 00.0000 Store at -20 C. CERTIFICATE OF ANALYSIS Long PCR Enzyme Mix is functionally tested in PCR amplification of 47.4 kb fragment from lambda
More informationChapter 8: Recombinant DNA. Ways this technology touches us. Overview. Genetic Engineering
Chapter 8 Recombinant DNA and Genetic Engineering Genetic manipulation Ways this technology touches us Criminal justice The Justice Project, started by law students to advocate for DNA testing of Death
More informationAppendix A. Introduction to PCR
Appendix A Introduction to PR In 1983, Kary Mullis at etus orporation developed the molecular biology technique that has since revolutionized genetic research, earning him the Nobel Prize in 1993. This
More informationBIO 121 LAB 10 - DNA I
BIO 121 LAB 10 - DNA I All cellular organisms store their hereditary information as the precise sequence of nucleotides in DNA, just as written information is stored as the precise sequence of letters
More informationRestorase DNA Polymerase with 10 Reaction Buffer. Catalog Number R1028 Storage Temperature 20 C TECHNICAL BULLETIN
Restorase DNA Polymerase with 10 Reaction Buffer Catalog Number R1028 Storage Temperature 20 C TECHNICAL BULLETIN Product Description Restorase DNA Polymerase with 10 Reaction Buffer combines Sigma s Long
More informationQuestions from chapters in the textbook that are relevant for the final exam
Questions from chapters in the textbook that are relevant for the final exam Chapter 9 Replication of DNA Question 1. Name the two substrates for DNA synthesis. Explain why each is necessary for DNA synthesis.
More informationFunctional Genomics Research Stream. Research Meeting: June 19, 2012 SYBR Green qpcr, Research Update
Functional Genomics Research Stream Research Meeting: June 19, 2012 SYBR Green qpcr, Research Update Updates Alternate Lab Meeting Fridays 11:30-1:00 WEL 4.224 Welcome to attend either one Lab Log thanks
More informationAccura High-Fidelity Polymerase
Accura High-Fidelity Polymerase FOR RESEARCH USE ONLY. NOT FOR HUMAN OR DIAGNOSTIC USE Lucigen Corporation 2905 Parmenter St, Middleton, WI 53562 USA Toll Free: (888) 575-9695 (608) 831-9011 FAX: (608)
More informationApplication Notes
Application Notes Superior Performance and Flexibility In order to minimize errors in DNA replication during PCR, it is essential to choose a high-fidelity DNA polymerase enzyme. The introduction of errors
More informationDETERMINATION OF THE Rh FACTOR BY PCR
DETERMINATION OF THE Rh FACTOR BY PCR Ref.: PCR2 1. EXPERIMENT OBJECTIVE The aim of this experiment is to introduce students to the principles and practice of the Polymerase Chain Reaction (PCR) by studying
More informationPCR PRIMER DESIGN SARIKA GARG SCHOOL OF BIOTECHNOLGY DEVI AHILYA UNIVERSITY INDORE INDIA
PCR PRIMER DESIGN SARIKA GARG SCHOOL OF BIOTECHNOLGY DEVI AHILYA UNIVERSITY INDORE-452017 INDIA BIOINFORMATICS Bioinformatics is considered as amalgam of biological sciences especially Biotechnology with
More informationThe Techniques of Molecular Biology: Forensic DNA Fingerprinting
The Techniques of Molecular Biology: Forensic DNA Fingerprinting Revised Fall 2017 Laboratory Safety: Lab coat, long pants, closed-toe shoes, safety goggles, and nitrile or latex gloves are required. Learning
More informationBlood direct 2x PCR Mastermix. Data sheet. Order No. BS reactions x 20 µl. (For research and in vitro applications only) Batch No.
Data sheet Order No. BS91.222.0250 250 reactions x 20 µl Order No. BS91.222.1250 1250 reactions x 20 µl (For research and in vitro applications only) Batch No.: Best before: Appearance: Colour: 1 Description
More informationAppendix A DNA and PCR in detail DNA: A Detailed Look
Appendix A DNA and PCR in detail DNA: A Detailed Look A DNA molecule is a long polymer consisting of four different components called nucleotides. It is the various combinations of these four bases or
More informationCERTIFICATE OF ANALYSIS. For. Red i-taq DNA Polymerase
CERTIFICATE OF ANALYSIS For Red i-taq DNA Polymerase Version 1 (110804) Catalog i-taq6 Size 500 units For Research Use Only and Not For Human and Animal Therapeutic and Diagnostic Use. Disclaimer: THE
More informationCalifornia Institute of Technology. Directed evolution. Dr. F.H. Arnold s lab
Directed evolution. Dr. F.H. Arnold s lab May 4, 1999 Mutagenic PCR -[Mn] The amount of Mn used in the reaction should be titrated to produce the desired mutagenic rate. Libraries that have close to 30%
More informationMolecular Genetics II - Genetic Engineering Course (Supplementary notes)
1 von 12 21.02.2015 15:13 Molecular Genetics II - Genetic Engineering Course (Supplementary notes) Figures showing examples of cdna synthesis (currently 11 figures) cdna is a DNA copy synthesized from
More informationOne Step SYBR PrimeScript RT-PCR Kit II (Perfect Real Time)
Cat. # RR086A For Research Use One Step SYBR PrimeScript RT-PCR Kit II Product Manual Table of Contents I. Description...3 II. III. IV. Principle...3 Components...5 Storage...6 V. Features...6 VI. VII.
More informationFeedback D. Incorrect! No, although this is a correct characteristic of RNA, this is not the best response to the questions.
Biochemistry - Problem Drill 23: RNA No. 1 of 10 1. Which of the following statements best describes the structural highlights of RNA? (A) RNA can be single or double stranded. (B) G-C pairs have 3 hydrogen
More informationExploring Genetic Variation in a Caffeine Metabolism gene LAB TWO: POLYMERASE CHAIN REACTION
Exploring Genetic Variation in a Caffeine Metabolism gene LAB TWO: POLYMERASE CHAIN REACTION Purpose: In this laboratory, we will set up a polymerase chain reaction to amplify the region of the caffeine
More informationInstructions for Use Life Science Kits & Assays
Instructions for Use Life Science Kits & Assays Content Content 1 Product and order number... I 2 Storage conditions... I 3 Description... II 3.1 Quality data... II 3.2 Unit definition... II 4 Delivered
More informationPCR-ReIated Products User's Instruction
ISO 9001:2000 Certified PCR-ReIated Products User's Instruction SBS Genetech Co.,Ltd. Table of Contents Cat. No. Product Name Page EUT-500 ER-500 EP-500 EQ2.2-100/2.5-100/5.2-100/5.5-100 EN-1/2 U-Taq DNA
More informationBio Rad PCR Song Lyrics
Bio Rad PCR Song Lyrics There was a time when to amplify DNA, You had to grow tons and tons of tiny cells. (Oooh) Then along came a guy named Dr. Kary Mullis, Said you can amplify in vitro just as well.
More informationThe most popular method for doing this is called the dideoxy method or Sanger method (named after its inventor, Frederick Sanger, who was awarded the
DNA Sequencing DNA sequencing is the determination of the precise sequence of nucleotides in a sample of DNA. The most popular method for doing this is called the dideoxy method or Sanger method (named
More informationAccuScript High Fidelity 1 st Strand cdna Synthesis Kit
AccuScript High Fidelity 1 st Strand cdna Synthesis Kit INSTRUCTION MANUAL Catalog #200820 Revision B.01 For In Vitro Use Only 200820-12 LIMITED PRODUCT WARRANTY This warranty limits our liability to replacement
More informationThe Polymerase Chain Reaction. Chapter 6: Background
The Polymerase Chain Reaction Chapter 6: Background PCR Amplify= Polymerase Chain Reaction (PCR) Invented in 1984 Applications Invention of PCR Kary Mullis Mile marker 46.58 in April of 1983 Pulled off
More informationB. Incorrect! Ligation is also a necessary step for cloning.
Genetics - Problem Drill 15: The Techniques in Molecular Genetics No. 1 of 10 1. Which of the following is not part of the normal process of cloning recombinant DNA in bacteria? (A) Restriction endonuclease
More informationReverse Transcriptase Reverse Transcriptase 100 µl 5X RT Buffer 0.1 M DTT 500 µl Storage -20 C for 24 months
www.smobio.com Product Information Reverse Transcriptase ExcelRT series RP1000 20,000 units Reverse Transcriptase 100 µl 5X RT Buffer 1 ml 0.1 M DTT 500 µl Storage -20 C for 24 months Description The ExcelRT
More informationPCR Introduction. Biochemistry Laboratory
PCR Introduction Biochemistry Laboratory 2010. 11 Mechanism of DNA Synthesis DNA polymerase extends the primer by sequentially adding a single dntp (datp, dgtp, dctp or dttp) that is complementary to the
More informationBasic lab techniques
Basic lab techniques Sandrine Dudoit Bioconductor short course Summer 2002 Copyright 2002, all rights reserved Lab techniques Basic lab techniques for nucleic acids Hybridization. Cut: restriction enzymes.
More informationBasic Steps of the DNA process
As time pasted technology has improve the methods of analyzing DNA. One of the first methods for the analysis of DNA is known as Restriction Fragment Length Polymorphism (RFLP). This technique analyzed
More informationGuidelines for Developing Robust and Reliable PCR Assays
Guidelines for Developing Robust and Reliable PCR Assays Leta Steffen, PhD Applications Scientist Promega Corporation Outline 1) PCR reaction components What is in the reaction? How does it affect assay
More informationPV92 PCR Bio Informatics
Purpose of PCR Chromosome 16 PV92 PV92 PCR Bio Informatics Alu insert, PV92 locus, chromosome 16 Introduce the polymerase chain reaction (PCR) technique Apply PCR to population genetics Directly measure
More informationBiotechnology. Chapter 20. Biology Eighth Edition Neil Campbell and Jane Reece. PowerPoint Lecture Presentations for
Chapter 20 Biotechnology PowerPoint Lecture Presentations for Biology Eighth Edition Neil Campbell and Jane Reece Lectures by Chris Romero, updated by Erin Barley with contributions from Joan Sharp Copyright
More informationUNCC Biotechnology and Bioinformatics Camp. Dr. Jennifer Weller Summer 2010
UNCC Biotechnology and Bioinformatics Camp Dr. Jennifer Weller Summer 2010 PCR Topic: the Polymerase Chain Reaction (PCR) 1988, Saiki, Mullis et al. proposed using a heat stable polymerase to carry out
More informationBootcamp: Molecular Biology Techniques and Interpretation
Bootcamp: Molecular Biology Techniques and Interpretation Bi8 Winter 2016 Today s outline Detecting and quantifying nucleic acids and proteins: Basic nucleic acid properties Hybridization PCR and Designing
More informationPCR Protocol Cooke Lab July 30, 2012
PCR Protocol Cooke Lab July 30, 2012 Contact Adriana Arango-Velez Contents 1. Before performing the PCR 2. Recommendations for the PCR 3. Performing the PCR 4. General Thermocycler program 5. Stock solutions
More informationPolymerase Chain Reaction (PCR) May 23, 2017
Polymerase Chain Reaction (PCR) May 23, 2017 Outline History of PCR Uses of PCR How PCR works How to set up and run PCR The structure of DNA PCR Polymerase chain reaction Selective amplification of target
More informationLecture 18. PCR Technology. Growing PCR Industry
Lecture 18 PCR Technology Growing PCR Industry Basic PCR, Cloning of PCR product, RT-PCR, RACE, Quantitative PCR, Multiplex PCR, Hot start PCR, Touchdown PCR,PCR sequencing.. How PCR started The DNA duplex
More informationMonsterScript Reverse Transcriptase
MonsterScript Reverse Transcriptase Cat. Nos. MSTA5110, and MSTA5124 Connect with Epicentre on our blog (epicentral.blogspot.com), Facebook (facebook.com/epicentrebio), and Twitter (@EpicentreBio). www.epicentre.com
More informationAssociate Professor Chatchawan Srisawat MD. Ph.D
POLYMERASE CHAIN REACTION Associate Professor Chatchawan Srisawat MD. Ph.D POLYMERASE CHAIN REACTION In vitro technique for amplification of the specified DNA sequences. It enables us to produce enormous
More informationDNA Technology. Asilomar Singer, Zinder, Brenner, Berg
DNA Technology Asilomar 1973. Singer, Zinder, Brenner, Berg DNA Technology The following are some of the most important molecular methods we will be using in this course. They will be used, among other
More informationFig. 16-7a. 5 end Hydrogen bond 3 end. 1 nm. 3.4 nm nm
Fig. 16-7a end Hydrogen bond end 1 nm 3.4 nm 0.34 nm (a) Key features of DNA structure end (b) Partial chemical structure end Fig. 16-8 Adenine (A) Thymine (T) Guanine (G) Cytosine (C) Concept 16.2: Many
More informationEnhanced Arginase production: rocf
Enhanced Arginase production: rocf Purpose and Justification: Bacillus subtilis produces urease, which catalyses the hydrolysis of urea into ammonium and carbonate. Since the cell wall of the bacteria
More informationEnzymes and Cellular Regulation
Why? Enzymes and Cellular Regulation What are the factors that regulate the rate at which enzymes catalyze reactions? Digestive enzymes are protein-based biological catalysts that play important roles
More informationQUICK-Clone TM User Manual. cdna
QUICK-Clone TM User Manual cdna PT1150-1 (PR752268) Published 25 May 2007 Table of Contents I. Introduction 3 II. Applications Discussion 4 A. Primer Design 4 B. Setting up the PCR Reaction 4 C. Example
More informationGenetic Fingerprinting
Genetic Fingerprinting Introduction DA fingerprinting In the R & D sector: -involved mostly in helping to identify inherited disorders. In forensics: -identification of possible suspects involved in offences.
More informationAffinityScript One-Step RT-PCR Kit
AffinityScript One-Step RT-PCR Kit INSTRUCTION MANUAL Catalog #600188 Revision A For In Vitro Use Only 600188-12 LIMITED PRODUCT WARRANTY This warranty limits our liability to replacement of this product.
More information3color RT HS-PCR Mix SYBR Ready-to-use mix for real-time Hot Start PCR with SYBR Green. Dedicated for white reaction tubes and plates.
3color RT HS-PCR Mix SYBR Ready-to-use mix for real-time Hot Start PCR with SYBR Green. Dedicated for white reaction tubes and plates. version 0217 250 reactions in 20 μl Cat. # 2000-250S 2500 reactions
More informationSensitivity vs Specificity
Viral Detection Animal Inoculation Culturing the Virus Definitive Length of time Serology Detecting antibodies to the infectious agent Detecting Viral Proteins Western Blot ELISA Detecting the Viral Genome
More informationCovalently bonded sugar-phosphate backbone with relatively strong bonds keeps the nucleotides in the backbone connected in the correct sequence.
Unit 14: DNA Replication Study Guide U7.1.1: DNA structure suggested a mechanism for DNA replication (Oxford Biology Course Companion page 347). 1. Outline the features of DNA structure that suggested
More information