Molecular Diagnostics: DNA Amplification Methods

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What is the name of the scientist who invented the polymerase chain reaction (PCR) method?

Kary B. Mullis (correct)

Buckingham

La Jolla

Taq polymerase

What is the primary function of the primers in PCR?

To denature the DNA template

To extend the binding site of the opposite primer

To bind to the complementary strand of the target fragment (correct)

To synthesize new strands of DNA

What is the purpose of heating the DNA template in PCR?

To extend the binding site of the primer

To cool the reaction mixture

To denature the DNA template (correct)

To promote primer annealing

What is the result of the PCR process?

Amplification of the target fragment Signup and view all the answers

What is the name of the enzyme used in PCR?

Taq polymerase Signup and view all the answers

What is the advantage of PCR technology?

It is a widely used method of nucleic acid amplification Signup and view all the answers

What is target amplification similar to?

Growing cells in culture and allowing them to multiply Signup and view all the answers

What is the advantage of target amplification over cell culture?

It takes minutes to hours to replicate nucleic acid Signup and view all the answers

What is the first and most used method for amplifying target nucleic acid?

PCR Signup and view all the answers

Who won the Nobel Prize in Chemistry in 1993?

Kary B. Mullis Signup and view all the answers

What will be discussed in this topic?

The components and variants of PCR Signup and view all the answers

What is the purpose of this topic?

To explore the applications of PCR and its variants Signup and view all the answers

What is the result of repeating the steps of denaturation, annealing, and extension in PCR?

An exponential increase in the total number of target DNA copies Signup and view all the answers

What is the temperature range for the annealing step in PCR?

50-70°C Signup and view all the answers

What is the purpose of the denaturation step in PCR?

To separate the DNA strands Signup and view all the answers

What is the optimal temperature range for DNA polymerase to act in PCR?

68-72°C Signup and view all the answers

What is an important parameter in PCR primer design?

GC content Signup and view all the answers

What is the formula for the number of PCR product copies?

2ˆN Signup and view all the answers

What is a major advantage of multiplex PCR over singleplex PCR?

Reduced reagent usage Signup and view all the answers

What is the primary purpose of using sequence-specific PCR?

To design primers with specific mismatch principles Signup and view all the answers

What is a common problem encountered during multiplex PCR?

Uneven amplification Signup and view all the answers

What is the primary application of multiplex PCR in a clinical molecular laboratory?

To detect different pathogens in a single tube Signup and view all the answers

Who first reported multiplex PCR in 1988?

Chamberlain et al. Signup and view all the answers

What type of PCR involves using RNA templates?

Reverse transcriptase PCR Signup and view all the answers

What is the main difference between real-time PCR and conventional PCR?

Amplification process detection Signup and view all the answers

What does the CT (threshold cycle) parameter represent?

The cycle number at which fluorescence exceeds the fixed threshold Signup and view all the answers

What is the advantage of real-time PCR quantitation?

Elimination of post-PCR processing Signup and view all the answers

What does the fluorescence threshold signify?

A fixed level significantly above the baseline fluorescence level Signup and view all the answers

What is the correlation between the starting copy number of the nucleic acid target and the fluorescence increase?

The higher the starting copy number, the sooner the fluorescence increase Signup and view all the answers

What is the purpose of setting a fixed fluorescence threshold?

To determine the CT parameter Signup and view all the answers