| 摘要 |
A method for determining the presence or absence of a variant nucleotide in a diagnostic region of a target nucleic acid sequence in a sample comprising a first primer and a blocking oligonucleotide probe wherein said first primer and said blocking probe overlap at a region that acts as an amplification priming region and the oligonucleotide probe further comprises a diagnostic target region binding sequence based on a reference sequence without a variant nucleotide, and a second downstream primer, carrying out an amplification reaction using the first and second primers with the oligonucleotide probe and subjecting the amplified products hybridised to the oligonucleotide probe to melting profile analysis to determine the presence of the variant nucleotides in the diagnostic region. Either of the first primer or the oligonucleotide probe can be labelled with FRET labels. The primers and probes are claimed. Alternatively claimed are allele specific primers for EGFR mutations. |
