Quantitation of specific transcripts by RT-PCR SNuPE assay.

The single nucleotide primer extension assay (SNuPE) provides a means of identifying and quantifying allelic variants in DNA and RNA. ~1'2) It has many uses, including the measurement of specific RNA levels, as well as the study of allelic variation and allele-specific expression. The SNuPE assay is done after amplification by PCR of a segment containing the allelic difference, which may be even a single nucleotide. The assay consists of the enzymatic extension by 1 base of a primer just 5' to the position of mismatch in the presence of a [32p]dNTP specific for either the wild-type or variant sequence. The ratio of radioactivity in the primer products of these two reactions is then determined after denaturing gel electrophoresis.