Polymerase Chain Reaction TAQ Polymerase, Primer, Nucleotide YouTube Lecture Handouts
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Polymerase Chain Reaction (PCR) : TAQ polymerase, Primer, Nucleotide - Covid-19 RT-PCR Test|GS
Title: Polymerase Chain Reaction
Where it is used –
- Crime investigation
- Genetic research
- Molecular biology
What is Polymerase?
What is Chain Reaction?
Polymerase Chain Reaction What is Required?
- Thermal cycler or PCR machine – reaction to occur –
- TAQ polymerase (Thermus Aquaticus – heat tolerant bacteria form where it was extracted) – enzyme to create copies of DNA – hot springs active at 70° Celsius
- Primer – polymerase cannot start reaction it can just propagate it – so required to start reaction – short sequence of nucleotide (20) – help select exact portion to amplify – 2 primers are used.
- DNA Template – segment to amplify
- Nucleotide – building block for DNA synthesis
Polymerase Chain Reaction Steps
- Denaturation - 96° Celsius – separate 2 DNA strands
- Annealing – heat and cool in metal – remove internal defects – raise temperature to 72° and then cool to 55° so that primer can bind to single stranded DNA – primers bind by complementary base pairing that is A to T and C to G
- Extension – free nucleotide to make new DNA – increase temperature to 72° and TAQ polymerase extend primer and form new strands
- Repeated heating and cooling for new DNA to be synthesised
- 4 new DNA – substrate will double we get 8 … and so it is a chain reaction
- Product of 1st reaction used as substrate for next DNA
- With few hundred reactions we get billion of copies of target DNA – again temperature lowered to 15° to store products
- To check if the products are correct – agarose gel electrophoresis is used
✍ Mayank