Conditions and Factors Affecting PCR
Factors Affecting PCR:
PCR Reaction Volume
If you are increasing the PCR reaction volume proportionally to a PCR reaction that is optimized or works well, in general you will increase the amount of product with increasing volume. Exceptions to this include older mineral oil PCR machines and PCR thermocyclers without a heated-lid. In these cases, increasing the PCR reaction volume will negatively affect the PCR product and efficiency of the reaction.
PCR Primer Design
Primer are usually chosen to be between 18-25 nucleotides in length. Longer primers also work 30-40 nucleotides or bp during pcr
A GC clamp at the 5' ends of the primers helps.
One can thus design primers with 1 or 2 nucleotides GC at the start and ends of the primers if possible