期刊
NEOPLASIA
卷 8, 期 2, 页码 153-162出版社
ELSEVIER SCIENCE INC
DOI: 10.1593/neo.05754
关键词
exponential RNA amplification; whole transcriptome amplification; prostate cancer; laser capture microdissection; FFPE tissue
类别
资金
- NCI NIH HHS [P50 CA069568, U01 CA111275, P50CA69568, UO1 CA111275-01] Funding Source: Medline
- NIA NIH HHS [R01 AG021404, R01AG21404] Funding Source: Medline
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.
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