Semi-quantitative RT-PCR for comparison of mRNAs in cells with different amounts of housekeeping gene transcripts

Mol Cell Probes. 1998 Oct;12(5):283-91. doi: 10.1006/mcpr.1998.0182.

Abstract

A simple method is reported here for the semi-quantitative assay of mRNAs in the presence of an exogenous mRNA (pBR322 transcript) as a standard. This method uses the co-reverse transcription and co-amplification (co RT-PCR) of the target and standard mRNAs. This procedure enables transcripts to be compared when the differentiation process affects the transcription pattern of the beta-actin housekeeping gene, a commonly used internal standard. This method is sensitive and avoids constructing internal competitive RNA standards. As an example, it is shown that beta-actin transcription decreases and type V adenylyl cyclase transcription increases in adipocytes, when compared to preadipocytes.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / genetics*
  • Adipocytes / cytology
  • Adipocytes / metabolism*
  • Animals
  • Cells, Cultured
  • DNA Primers
  • DNA, Complementary
  • Epididymis
  • Male
  • Plasmids
  • Quality Control
  • RNA, Messenger / analysis*
  • RNA, Messenger / genetics*
  • Rats
  • Rats, Sprague-Dawley
  • Reproducibility of Results
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • Reverse Transcriptase Polymerase Chain Reaction / standards
  • Transcription, Genetic*

Substances

  • Actins
  • DNA Primers
  • DNA, Complementary
  • RNA, Messenger