A solution hybridization method for quantification of mRNAs: Determining the amount and stability of oncogene mRNA

Abstract

A solution hybridization method for the quantification of specific mRNAs is described. This assay utilizes complementary RNA probes prepared by in vitro transcription, sandwich hybridization in solution, and affinity-based hybrid collection. The possibility of using this method for crude biological samples without purifying mRNAs makes it ideal when accurate quantification of multiple samples is needed. Human N-myc oncogene transcript was used as a model and as little as 0.24 pg (2 × 10⁵ molecules) of N-myc mRNA could be detected. Using this assay it was shown that human neuroblastoma IMR-32 cells contain ′500 N-myc mRNA molecules per cell having a half-life of ′35 min.