Q7 7.You are tasked with amplifying a fragment of interest from a sample of genomic DNA by usingPCR.List the components that need to be included in any standard PCR, give the function of each.The figure below represents a step during the first PCR cycle. Indicate the polarities of bothDNA strands and the primers. Use arrows to indicate the direction of synthesis of the newstrands.PrimerPrimerc)You want to PCR amplify only the DNA region indicated in the block. Design a suitable forwardand reverse primer each 10 bp in length, give the primers sequences 5' – 3'.5'...GCTAGTCGGAAGTCGCCAGTGCA..3'...CGATCAGCCTTCAGCGGTCACGT.CATAGACGTTACCAGTATGGCGA... 3'GTATCTGCAATGGTCATACCGCT... 5'd)Can RNA be used as the template for a PCR reaction? Motivate your answer.

PCR stands for the polymerase chain reaction. It is described as a test that is used to detect the…

Answered: 7. You are tasked with amplifying a…

Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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Similar questions

Explain why DNA ladders are usually included during gel electrophoresis. One aspect of PCR that can be modified is the annealing temperature. In general, higher annealing temperatures show more specificity towards a single template, whereas lower annealing temperatures show less specificity and may bind to multiple regions throughout the genome. Discuss how using an annealing temperature that is too high or too low might influence the results of a PCR assay (and gel electrophoresis results) such as the one used in this study.
Examine the DNA fragment sequence below. Your job is to design primers for PCR that would be able to amplify this DNA fragment. Design the primers so that they are 7 bases in length. Don’t forget to indicate direction (polarity) of the primers. Also describe where the primer would bind (i.e. top or bottom strand, left or right side of the DNA strand). Please organize your response so that each primer, and associated information, is separated by at least one blank line 5’ - TCCACTTGCTGTGTAGCTAAATCATATAACAG3’ - AGGTGAACGACACATCGATTTAGTATATTGAC
This is what the LAB FLOW would look like: CULTURE BACTERIA EXTRACT RNA RANDOM HEXAMER REVERSE TRANSCRIPTION OF RNA PCR OF lacZ GENE ELECTROPHORESIS If you wanted to prepare only one PCR reaction, how much of each reagent would you add to the PCR tube? What would be the final primer concentration if 0.5 μl of 10 μM primers were added to a PCR reaction with a final volume of 20 μl?
The process of PCR essentially revolves around three phases. Briefly describe these phases and the events that occur in them. Take note the temperature on which these phases take place.
1. Why is each of the following reagents required to be in the PCR reaction tube? Primers, Taq DNA polymerase, nucleotide mix, reaction buffer 2. Imagine that you forgot to add nucleotides to your PCR master mix. How would this affect the outcome of your PCR reactions? a) If you made the mistake described in the previous question, which control would give you unexpected results? (In other words, if you forgot to add nucleotides, which control reaction tube would have results that are different that they would be if you had prepared them correctly?) 3. If you found that there was DNA amplification in your negative control tube, what could be an explanation for that result?
A student carries out PCR using the following steps: step 1: 94C for 1 minuteStep 2: 60C for 30seconds Step 3: 72C for 30 seconds The student is examining a 1000-bp DNA fragment as part of their research project. If the limit of detection of this molecule 9 x 108 molecules, what is the minimum number of PCR cycles you would have to run to detect PCR product generated from a single molecule of the template? 2. Two double-stranded fragments of DNA are exactly the same length. At 89 C, fragment A has completely denatured which means the two strands have separated. At that temperature, fragment B is still double-stranded. How might these fragments differ to result in different denaturation temperatures?
There are many types of PCR techniques available nowadays. Explain the concept of nested PCR and real-time PCR.
1. You decide to perform dideoxy sequencing on a PCR product. You add the appropriate 32P- labeled primer, DNA polymerase, DNA template (the PCR product), buffer, DNTP mix, and a small amount of one of the four ddNTPs to four reaction tubes. You run the reactions in the thermal cycler, load each reaction into a separate lane of a polyacrylamide gel, and separate the products by gel electrophoresis. In the figure below, the lanes are labeled according to the ddNTP added. ddATP ddTTP ddCTP ddGTP - Lane 1 2 3 4 a) In lane 5, draw what you would expect to see if you prepared a reaction using a nucleotide mix containing only DATP, DTTP, dCIP, and dGTP. b) In lane 6, draw what you would expect to see if you prepared a reaction using a nucleotide mix containing only ddATP, DTTP, dCIP, dGTP. ||
Look at each PCR component listed below. For each one, determine which steps(s) of the PCR reaction (denaturation, annealing or extension) would be directly affect if that component were missing. Taq polymerase: Oligonucleotide primers: DNA template: Deoxynucleotides (A, T, G and C): Imagine that you correctly prepare your PCR reaction mixture, but there is something wrong with the thermal cycler. Describe what would happen if: The thermal cycler was stuck on 940C: The thermal cycler cycled between 600C and 720C, but never reached 940C The thermal cycler cycled between 940C and 720C, but never reached 600
PCR primers Below is a 300 base pair fragment of DNA. The top strand is written in the 5' to 3' direction. The bottom strand is written 3' to 5'. There are also two primer sequences; both primers are written 5' to 3'. Note that we are displaying a double-stranded DNA fragment, but primers will only bind to one of the two displayed strands. 5' ACCGȚAGCTATATGCTATCGTGACGTATCGGCGCATTAAȚCGGGATCGAT 3 50 3' TGGCÁTCGATATACOATAGCACTOCATAGCCGCGTAATTÀGCCCTAGCTÀ 5' 5' AGCTÇGCTAGCAGGAGAGAȚATCGÇTCATAGCTCCGATCGATGCCGCTAA 3 3' TCGAGCG ATCGTCCTCTCTÁTAGCGAGTATCGAGÓCTAGCTACGGCGATİ 5' 100 5' TATAGCTCTÇTGCGGATATÇGCATATACCẠ AGGCCCTACGTATGTAGCTA 3 150 3' ATATČGAGAGACOCCTATAGCGTATATGGTTCCGGGATGČATACATCGAŤ 5' 5 TGCGTATATÇGGAGAGTCCTGGATATGGAGCTTGACTGCAGAGAGCTCGA 3 200 3' ACGCÁTATAGCCTCICAGGÁCCTATACCTCGAACTGACGTCTCTCGAGCT 5' 5' TATGCGCTTAGGCCGTATATGCTTGGGGAAAGCTCTATGTATGCTATGTG 3 3. ATACGCGAATCCGGCATATACGAACCCCTÍTCGAGATACATACGATACAC 5' 250 5' TGCATGTGCTATGCAACGTTCOGATTGCGȚAGCAGTAATAGCGCCGATTG 3 300 3'…
The polymerase chain reaction (PCR) is used by scientists to amplify DNA, particularly when the quantity of DNA is very small, mixed, or contaminated with other organisms. Explain (with words) the how PCR works using a diagram to help illustrate this (show a minimum of three cycles to illustrate your point).
1. Explain what primers are and what purpose they serve in a PCR reaction. Explain the main steps and temperatures of one PCR cycle. 2. Explain the purposes of the following components of gel electrophoresis: agarose, loading buffer, DNA ladder/marker, electrode buffer, electrodes. 3. Explain the expected results of the Alu sequence at PV92 that we tested, in terms of numbers of bands and their sizes. A labeled drawing would suffice.

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