A sensitive quantification method for evaluating the level of hepatocyte growth factor and c-met/HGF receptor mRNAs in the nervous system using competitive RT-PCR

Woong Sun, Hiroshi Funakoshi, Kunio Matsumoto, Toshikazu Nakamura

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Conventional RNA quantification methods such as Northern blots or RNase protection assays are often not sufficiently sensitive to measure mRNA levels in a small neuronal region. The reverse transcription-polymerase chain reaction (RT-PCR) is a sensitive alternative that can be used to determine the relative amount of mRNAs in tissues or cells. However, this method does not directly yield the absolute value of mRNA abundance because of the exponential nature of PCR. Using synthetic RNA competitors, we developed a competitive RT-PCR to evaluate the absolute amount of hepatocyte growth factor (HGF) and c-met/HGF receptor mRNAs in neural tissues. Here we describe the procedures we used to measure HGF and c-met mRNA levels in the punched ventral horn of the mouse spinal cord. This protocol provides a rapid, sensitive and accurate means of measuring mRNA levels and allows for comparison of the expression of related genes at one time and in a tiny piece of sample from a specific neuronal region. Themes: Development and regeneration. Topics: Neurotrophic factors: expression and regulation. Copyright (C) 2000 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)190-197
Number of pages8
JournalBrain Research Protocols
Volume5
Issue number2
DOIs
Publication statusPublished - 2000 Apr 1
Externally publishedYes

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Keywords

  • c-met
  • Competitive RT-PCR
  • HGF
  • mRNA
  • Quantification
  • Spinal cord

ASJC Scopus subject areas

  • Neuroscience(all)

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