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Single nucleotide primer extension: quantitative range, variability, and multiplex analysis.

A D Greenwood·D T Burke
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Abstract

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

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AllelesAnimalsBase SequenceDNA PrimersHypoxanthine PhosphoribosyltransferaseMenkes Kinky Hair SyndromeMiceMice, Inbred C57BLMolecular Sequence DataOrnithine CarbamoyltransferasePoly TPolymerase Chain ReactionPolymorphism, GeneticRibonucleasesSensitivity and SpecificitySpecies SpecificityTranscription, GeneticJournal ArticleResearch Support, Non-U.S. Gov'tResearch Support, U.S. Gov't, P.H.S.

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A D Greenwood, D T Burke. 1996. Single nucleotide primer extension: quantitative range, variability, and multiplex analysis.. https://doi.org/10.1101/gr.6.4.336

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