Effect of Reagent Design on Mutation Assay Performance in Cell Line RNA or DNA.

aΔCT is the difference in CT obtained from wild type and mutant templates in the allele-specific mutation assay.
bAssay names ending in “.1” were designed with the forward primer to be specific for the mutant sequence and assay names ending in “.2” were designed with the reverse primer specific for the mutant sequence.
cΔCT obtained when an allele-specific primer with Tm about 60° was used without a blocker.
dΔCT obtained when a blocker was used in combination with an allele-specific primer.
eΔCT obtained when a low Tm allele-specific primer was used.
fΔCT obtained when both a blocker and low Tm allele-specific primer were used in combination.
gSynthetic mutant DNA was used for Assay Mut6.1 due to the unavailability of cell lines carrying this mutation. Wild type DNA was obtained from HeLa cells.