INTERPRETATION OF FLU-1, FLU-2, AND K-VARIANT MUTATIONS BY COP-PCR
Interpretation of genotype at the flu-1 and flu-2 loci by COP-PCR was straightforward, and from our patient population, four heterozygous/affected patients were identified.
Adjustment of pH and [Mg.sup.2+] concentrations did not improve the specificity (not shown), and because other strategies substantially increased the number of steps and turnaround time of COP-PCR, the interpretation of Kvariant was based on POS-hex reaction products alone (NEG-hex reaction products were not used).
At first, the dibucaine and sil-1 mutations were also analyzed by COP-PCR (with DNA sequencing confirmation of affected individuals), but it became more expeditious for us to proceed directly to sequencing analysis, given the high frequency of dibucaine abnormalities in our population.
The COP-PCRs for the three mutations were performed simultaneously in a PC-960G gradient thermal cycler (Corbett Research), all with the same cycling conditions.