SINGLE NUCLEOTIDE PRIMER EXTENSION - QUANTITATIVE RANGE, VARIABILITY,AND MULTIPLEX ANALYSIS

The quantitative measurement of transcription products from homologous alleles at a diploid locus has broad application for the study of mam malian gene expression. Single nucleotide primer extension (SNuPE) ana lysis is a simple and sensitive method for allelic transcript discrimi nation requiring only 1 bp difference between alleles. In this study t he effective range, experimental variation, and the influences of poly (dT)-primed and gene-specific reverse transcriptions are characterized . The ability to analyze several genes fron 1 a single reverse transcr iption reaction is assessed as well. For the genes examined, the maxim um range of detection is reached when the minor transcript represents 1/250 of the major allele. Relatively little error is seen within or b etween assays and linearity of response is maintained over an approxim ately thousandfold range.