Polymerase Chain Reaction

What is the polymerase chain reaction (PCR)?

A method for making multiple copies of specific DNA fragments in vitro through enzymatic synthesis.

Who conceived the polymerase chain reaction (PCR)?

Kary B. Mullis conceived the PCR during a drive in April 1983.

List the three major steps of PCR.

1. Denaturation 2. Annealing 3. Extension

At what temperature does denaturation occur in PCR?

Denaturation occurs at 94-95°C.

What is the purpose of primers in PCR?

Primers bind to the DNA template to provide an initiation site for DNA synthesis.

What is Taq polymerase and why is it significant?

Taq polymerase is a heat-stable DNA polymerase used in PCR that can withstand the high temperatures of denaturation.

What can be a potential issue if too much template DNA is used in PCR?

Using too much template DNA can result in poor amplification due to saturation.

How is quantitative PCR (Q-PCR) different from standard PCR?

Q-PCR allows for the quantification of DNA while monitoring the PCR reaction in real time.

What is reverse transcription PCR (RT-PCR)?

RT-PCR is a technique that converts RNA into complementary DNA (cDNA) before amplification.

What role does magnesium chloride play in PCR?

Magnesium chloride acts as a cofactor necessary for the activity of Taq polymerase and affects DNA binding.

What is multiplex PCR?

Multiplex PCR involves using multiple sets of primers to simultaneously amplify different regions of DNA.

Explain the concept of primer design.

Primer design is the process of creating primers that are specific to the target sequence with optimal length and GC content.

What is a negative control in PCR?

A negative control is a sample without DNA to ensure any amplification observed comes from the intended template.

What can cause contamination in PCR?

Contamination can arise from the person setting up the reaction, cross-contamination of samples, and contaminated reagents.

Describe the purpose of agarose gel electrophoresis in PCR.

Agarose gel electrophoresis is used to separate and visualize PCR products by size.

What is digital PCR?

Digital PCR is a method that partitions a sample into many parallel PCR reactions for more accurate quantification.