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RT-PCR

Updated: Aug 20, 2020




Friends, I trust you have come across this three letters word PCR or RT-PCR many times since COVID-19 pandemic started.

PCR stands with Polymerase Chain Reaction

RT-PCR: Reverse Transcription – Polymerase Chain Reaction

What is PCR?

PCR (Polymerase Chain Reaction) – The PCR is a DNA amplification (increasing amount) system which produce a large amount of DNA in vitro (in test tube) from a very small sample of DNA and amplify it to a large enough amount to study in detail.

The procedure of PCR involves four main stages which are listed below:

(i) Denaturation – On heating DNA at 94 degree Celsius, DNA disassociate with single stranded DNA.

(ii) Primer Annealing – The temperature is reduced to 50-60 degree Celsius and primer is attached to targeted DNA.

(iii) DNA Synthesis – This is formation of DNA

(iv) Detection of Amplified products

Types of PCR: (i) RT-PCR (ii) Nested PCR (iii) Multiplex PCR

(i) RT-PCR (Reverse Transcription PCR) – RT-PCR was developed to amplify RNA, in the technique in RNA instead of DNA. In this process complementary DNA (cDNA) is first produced from RNA with the help of enzyme reverse transcriptase and them cDNA is amplified by PCR. Don’t confused with Real time RT–PCR, RT always stands with Reverse Transcriptase, Real Time is added because this process is real time conversion of RNA to DNA. RT-PCR is one of the most widely used laboratory methods for detecting the COVID-19 virus, Ebola virus and Zika virus.

(ii) Nested PCR – nested PCR was developed to increase the sensitivity and specificity of PCR technique.

(iii) Multiplex PCR – Two or more primer sets are used.

Application of PCR

· Genetic disease

· Neoplastic disease

· Forensic investigation

· Clinical microbiology – for diagnosis of various infectious diseases.

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