Question

Explain the polymerase chain reaction and include all starting materials required, what happens in each cycle, and how a scientific investigation might benefit from PCR.

Answer 1

Answer: Polymerase chain reaction(PCR) is a technique which is used to makenumber of copies of a particular segment of DNA,in a quick manner and accurately.It is a technique of purification or cloning.

PCR technique requires following materials for the process. Attached PCR technique diagram.

• Target sample,this is the biological sample where amplification is to be done).
• A Primer,these are short strands of DNA which adhere to target segment).
• Taq polymerase,the enzyme which takes care of replicating DNA.
• Nucleotides-nucleotides are dNTPs which are to added so DNA polymerase proceeds with the process.
• Buffer-100mM Tris-HCL buffer,Ph 9.0 to be maintained,15mM MgCl2,500mM KCL.
• PCR technique involves three steps which are as follows:
• Denaturation: In this step,the two stranded DNA is separated in to single stranded DNA.The DNA strand is heated to temperature of about 95 degree celsius.Each strand acts as a template where new strand is built.
• Annealing: In this step,temperature reduced and maintained between 40-70 degree celsius.Here the primer is added,by decreasing temperature,hydrogen bonds reforms and complimentary strand hybridized.so the primer gets attached to the DNA which is to be amplified and helps it to get ready for replication.
• Elongation(or)Extension:This process carried out at a temperature of 72degree celsius known as elongation temperature.At 72 degree celsius,Taq polymerase gets binded to single strandedDNA and starts replication using deoxy ribonucleoside triphosphates present in the reaction mixture.
• Applications: PCR mainly helps in acellular cloning,used to detect genetic diseases,detection of infectious diseases,identify different species by genetic finger printing etc.,

PCR Components PCR Process (One Cycle) 95°C - Strands Separate 1. Denaturing DNA Sample Primers Nucleotides HTT 55°C - Primers Bind Template 2. Annealing Taq Polymerase Mix Buffer PCR Tube ih 72°C - Synthesise New Strand 3. Extension PCR Cycle Thermal Cycler