Question

You are a leading scientist heading a research laboratory at SWOSU. Your student shows you a DNA gel of a PCR experiment that he recently conducted. To trouble shoot his experiment, you ask him about the ingredients he had in each PCR tube (4 pt)

Tube 1. Template DNA, Primers, DNA polymerase, nucleotides and buffer

Tube 2. Template DNA, Primers, DNA polymerase and buffer

Tube 3. Primers, DNA polymerase and buffer

Tube 4. Template DNA, Primers, Nucleotides and buffer

One of these tubes will have successful amplification of target DNA and others will not have any amplification.  In the space provided below explain the reason why there will be amplification one of the tubes while there is be no amplification in other tubes. (Do not just answer yes or no. Provide detailed explanation for each tube and provide justification for how the left-out component will impact the PCR process)

 

Tube 1._______________________________________________

 

Tube 2. ______________________________________________

 

Tube 3. ______________________________________________

 

Tube 4. ______________________________________________

Answer 1

Note: Here I am considering your buffer also contain MgCl2

Tube 1: Amplification will be observe in this tube as all the essential components of PCR reaction are present.

Tube 2: Nucleotides (dNTPs) is missing in this tube. No amplification will be observe in this tube. Because dNTPs are the buliding block for amplification of DNA, enzyme DNA polymerase cannot extend primer without dNTPs.

Tube 3: Again this tube is also missing dNTPs. In addition to nucleotides, no template is present in reaction mixture. Without template primer cannot bind to its target area. And therfore no amplification.

Tube 4: This tube is missing DNA polymerase. This is an enzyme which carry out amplification. It add dNTPs by reading template DNA. Without DNA polymerase no amplification will be observe on gel.