Question

Question 1

Let’s start with a relatively straightforward one – and make it topical.

1. Briefly explain what qPCR is and why it is different than traditional PCR. Address what the q stands for and why the technique is sometimes called real-time pcr.

Unfortunately – that real-time moniker became a problem because some people shortened it to RT-PCR.However, most usage of RT-PCR is something distinct from qPCR.

2. Describe that something else RT-PCR.

You can combine qPCR and RT-PCR – producing RT-qPCR. One application of RT-qPCR is detection of certain viruses – such as COVID-19.

3. For this application, describe why RT-qPCR is preferable to qPCR.
4. For this application, describe why RT-qPCR is preferable to RT-PCR.

Answer 1

Answer A

The qPCR is a technique in molecular biology that monitors DNA amplification during the PCR process is carried out and not at the end of the PCR completion i.e like in conventional PCR.

Tradition PCR or conventional PCR shows the results after loading the sample on an agarose gel and after the entire cycle is over but in real-time PCR we can monitor the amplification per cycle and we will know the success of PCR before completion of entire cycles.

q in qPCR stands for quantitative hence it is called qPCR in short.

In qPCR the real-time monitoring of amplification is noted hence qPCR is also called as real-time PCR.

Answer B

There are mainly 3 types of PCR - qPCR/real-time PCR, conventional PCR and Reverse Transcriptase PCR(RT PCR). Hence real-time PCR is not RT PCR but Reverse Transcriptase PCR is an RT PCR.

Reverse transcriptase PCR (RT-PCR) was initially developed to amplify RNA targets for viruses like HIV, HCV, and influenza it can also be used for COVID-19. In this method RNA is isolated and complementary DNA (cDNA) is synthesized by Reverse transcriptase PCR, qPCR can be later used to amplify the complementary DNA. Hence qPCR -RT PCR can be combined. qPCR -RT PCR is preferable for viruses as it can do a reverse transcription of viral RNA but the only qPCR will only amplify and have no use in the viral study.

Answer C

In the case of various Virus, the RNA has the genetic material and the absence of DNA is observed due to this detection is not possible. Hence RNA is isolated and complementary DNA (cDNA) is synthesized by Reverse transcriptase PCR, qPCR can be later used to amplify the complementary DNA. Hence qPCR -RT PCR can be combined.

Answer D

qPCR -RT PCR is preferable because it can make complementary DNA as well as amplify the DNA with real-time monitoring. In RT PCR only complementary DNA will be synthesized and amplification for the specific target primer will not be done.

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