A faithful method for PCR-mediated global mRNA amplification and its integration into microarray analysis on laser-captured cells

Kazuhiko Aoyagi, Takeshi Tatsuta, Michiko Nishigaki, Shingo Akimoto, Chikako Tanabe, Yoko Omoto, Shin ichi Hayashi, Hiromi Sakamoto, Michiie Sakamoto, Teruhiko Yoshida, Masaaki Terada, Hiroki Sasaki

研究成果: Article査読

80 被引用数 (Scopus)

抄録

Quantitative and qualitative analyses of mRNAs from a small number of cells are extremely important for studies on gene expression in various physiological and pathological conditions in multicellular organisms. We present here an effective method for high-fidelity global mRNA amplification for in vivo gene expression profiling of as few as 100 cells obtained by laser-captured microdissection (LCM). This method, called TALPAT, is based on T7 RNA polymerase-mediated transcription, adaptor ligation, and PCR amplification followed by T7-transcription. More than 80% of genes were commonly identified as a more than 3-fold changed gene among three gastric cancer cell lines using cRNA amplified by both TALPAT and the ordinary in vitro T7-transcription. The reproducibility of TALPAT was validated by microarray analysis on 100 breast cancer cells obtained by LCM. For the application of the LCM-TALPAT method, we successfully obtained expression profiles of gastric cancer cells and the mesenchymal cells, enabling us to understand in vivo cell-to-cell cross-talk in the microenvironment.

本文言語English
ページ(範囲)915-920
ページ数6
ジャーナルBiochemical and biophysical research communications
300
4
DOI
出版ステータスPublished - 2003 1月 24
外部発表はい

ASJC Scopus subject areas

  • 生物理学
  • 生化学
  • 分子生物学
  • 細胞生物学

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