Whole transcriptome amplification for gene expression profiling and development of molecular archives

Scott A. Tomlins, Rohit Mehra, Daniel R. Rhodes, Rajal B. Shah, Mark A. Rubin, Eric Bruening, Vladimir Makarov, Arul M. Chinnaiyan

Research output: Contribution to journalArticle

36 Scopus citations

Abstract

Expression profiling of clinically obtainable tumor specimens has been hindered by the need for microgram quantities of RNA. In vitro transcription (IVT)-based amplifications are most commonly used to amplify small quantities of RNA for microarray analysis. However, significant drawbacks exist with IVT-based amplification, and the need for alternative amplification methods remains. Herein, we validate whole transcriptome amplification (WTA), an exponential amplification technique that produces cDNA libraries and amplified target in 3 to 4 hours from nanogram quantities of total RNA using a combination of cDNA microarrays and quantitative polymerase chain reaction (PCR). We demonstrate that WTA material can serve as a "molecular archive" because a WTA cDNA library can be faithfully amplified through multiple rounds of PCR amplification, allowing it to serve as a bankable and distributable resource. To demonstrate applicability, WTA was combined with laser capture microdissection to profile frozen prostate tissues. Unlike most IVT-based and exponential amplification techniques, WTA does not depend on the presence of a poly-A tail. Thus, we demonstrate that WTA is compatible with artificially degraded RNA and RNA isolated from formalin-fixed paraffin-embedded tissues. Taken together, WTA represents a versatile approach to profile and archive cDNA from minute tumor samples and is compatible with partially degraded RNA.

Original languageEnglish (US)
Pages (from-to)153-162
Number of pages10
JournalNeoplasia
Volume8
Issue number2
DOIs
StatePublished - Feb 2006
Externally publishedYes

Keywords

  • Exponential RNA amplification
  • FFPE tissue
  • Laser capture microdissection
  • Prostate cancer
  • Whole transcriptome amplification

ASJC Scopus subject areas

  • Cancer Research

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  • Cite this

    Tomlins, S. A., Mehra, R., Rhodes, D. R., Shah, R. B., Rubin, M. A., Bruening, E., Makarov, V., & Chinnaiyan, A. M. (2006). Whole transcriptome amplification for gene expression profiling and development of molecular archives. Neoplasia, 8(2), 153-162. https://doi.org/10.1593/neo.05754