TRUPCR® Coagulation Panel Kit.
The TRUPCR® Coagulation Panel Kit is designed for the qualitative detection of Factor-II Mutation (Prothrombin, G20210A), Factor-V Leiden Mutation (G1691A) and MTHFR Mutation (C677T and A1298C) in human blood.
TRUPCR® Coagulation Panel Kit is an allelic discrimination real-time polymerase chain reaction (PCR) assay for qualitative detection of the mutation against a background of wild-type genomic DNA. In an allelic discrimination, two different probes specific for each allele are included in the PCR assay. Each probe is labelled with a different fluorescent dye (such as FAM or HEX/VIC) at its 5' end and contains a non fluorescent quencher at the 3' end. During qPCR amplification of the target DNA the probes will compete for binding across the variant region. The probe that is 100% homologous to the DNA binding site will preferentially bind and give a fluorescent signal as PCR proceeds. It follows that the wild-type sequence will give a strong amplification plot through one channel whilst giving a very weak signal through the alternative channel. Homozygous variant samples will give an exactly inverse result. A mismatch between probe and target greatly reduces the efficiency of probe hybridization and cleavage and no reporter dye is released. Thus, substantial increase in FAM or HEX/VIC dye fluorescence indicates homozygosity for the FAM- or HEX/VIC- specific allele. An increase in both signals indicates heterozygosity.