A method for the quantitative analysis of human heat shock gene expression using a multiplex RT-PCR assay

San Ming Wang, Janardan D. Khandekar, Karen L. Kaul, David J. Winchester, Richard I. Morimoto

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

A quantitative multiplex RT-PCR assay is described to measure the levels of messenger RNAs for eight human genes encoding the heat shock proteins (HSP) and molecular chaperones hsp90α, hsp90β, hsp70, hsc70, mtHsp75, Grp78 (BiP), hsp60 and hsp27. The basis of this assay is reverse transcription of total RNA isolated from human cells followed by amplification with PCR. By the careful selection of pairs of oligonucleotide primers corresponding to unique regions of each heat shock gene, selectivity can be attained such that messenger RNAs of multiple heat shock genes can be analyzed simultaneously in a single reaction. This method provides both the absolute and relative levels of each heat shock message by including in the reaction, reference control RNAs corresponding to in vitro transcripts of heat shock gene plasmids carrying small internal deletions.

Original languageEnglish (US)
Pages (from-to)153-161
Number of pages9
JournalCell Stress and Chaperones
Volume4
Issue number3
DOIs
StatePublished - Sep 1 1999

Fingerprint

Multiplex Polymerase Chain Reaction
Gene expression
Shock
Assays
Hot Temperature
Gene Expression
Genes
Chemical analysis
RNA
Messenger RNA
Molecular Chaperones
Gene encoding
DNA Primers
Transcription
Heat-Shock Proteins
Reverse Transcription
Amplification
Plasmids
Cells
Polymerase Chain Reaction

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

Cite this

A method for the quantitative analysis of human heat shock gene expression using a multiplex RT-PCR assay. / Wang, San Ming; Khandekar, Janardan D.; Kaul, Karen L.; Winchester, David J.; Morimoto, Richard I.

In: Cell Stress and Chaperones, Vol. 4, No. 3, 01.09.1999, p. 153-161.

Research output: Contribution to journalArticle

Wang, San Ming ; Khandekar, Janardan D. ; Kaul, Karen L. ; Winchester, David J. ; Morimoto, Richard I. / A method for the quantitative analysis of human heat shock gene expression using a multiplex RT-PCR assay. In: Cell Stress and Chaperones. 1999 ; Vol. 4, No. 3. pp. 153-161.
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