Mark Behlke, Chief Scientific OfficerIntegrated DNA Technologies AbstractIntegrated DNA Technologies (IDT) has developed a PCR system with improved specificity called rhPCR. The method employs modified primers that have an internal RNA residue and are 3’-end-blocked so as to be non-functional in the intact state. Cleavage by RNase H2 removes the 3’-block and the RNA base, activating the primers. Cleavage requires duplex formation of primer and target; mismatches near the cleavage site decrease the efficiency of cleavage by RNase H2, thereby increasing the specificity of the assay. The new assay shows markedly reduced potential for primer-dimer formation, enabling higher levels of multiplexing in single tube assays. rhPCR shows much higher mismatch discrimination than traditional allele-specific PCR and has utility in genotyping as well as in rare-allele detection.
Genomics