What is the role of salt in PCR reactions?
What is the role of salt in PCR reactions?
Successful PCR requires that the DNA duplex separates during the denaturation step and that primers anneal to the denatured DNA. Salt neutralizes the negative charges on the phosphate backbone of DNA, stabilizing double-stranded DNA by offsetting negative charges that would otherwise repel one another. Potassium chloride (KCl) is normally used in PCR amplifications at a final concentration of 50 mM. To improve amplification of DNA fragments, especially fragments between 100 and 1,000 bp, a KCl concentration of 70–100 mM is recommended. For amplification of longer products, a lower salt concentration appears to be more effective, whereas amplification of shorter products occurs optimally with higher salt concentrations. This effect is likely because high salt concentration preferentially permits denaturation of short DNA molecules over long DNA molecules.
It is important to note that a salt concentration above 50 mM can inhibit Taq polymerases.