Question

What enables PCR to amplify just one location (locus) in a person's genome as opposed to several loci scattered throughout the genome (what gives PCR its specificity)?

Answer 1

Ans. Uniqueness of Primers

Every gene locus at its own unique nucleotide sequence. The type of primer present in the PCR reaction mixture determines which locus is to be amplified.

Greater is the length of the primer, greater is its uniqueness i.e. lesser is the probability of finding multiple copies of the sequences complementary to target DNA on any locus. For example, a 5 nucleotides long primer may complementarily bind DNA every (1/4)⁵ = 1025 bp; a 10 nucleotides long may bind complementary DNA every (¼)¹⁰ = 1048576 bp. In PCR, the primer length is generally around 20 nucleotides (it may differ from lab to lab, protocol to protocol). The primer may complementarily bind the target DNA every 1099511627776 (=1/4)²⁰ i.e. 1.09 x 10¹³ base pair. So, the probability that the primer binds exclusively the target DNA to be amplified is almost 1 (human genome size= 3 x 10⁹ bp).

Thus, the uniqueness of primer ensures that only the target DNA is complementarily bound with the primers and the DNBA-primer complex is further elongated by Taq polymerases. The resultant product exclusively contains the target DNA only.