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

Research output: Contribution to journalArticle

80 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)915-920
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume300
Issue number4
DOIs
Publication statusPublished - 2003 Jan 24
Externally publishedYes

Keywords

  • Breast cancer
  • Expression profiling
  • Gastric cancer
  • Global mRNA amplification
  • LCM
  • Microarray

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint Dive into the research topics of 'A faithful method for PCR-mediated global mRNA amplification and its integration into microarray analysis on laser-captured cells'. Together they form a unique fingerprint.

  • Cite this

    Aoyagi, K., Tatsuta, T., Nishigaki, M., Akimoto, S., Tanabe, C., Omoto, Y., Hayashi, S. I., Sakamoto, H., Sakamoto, M., Yoshida, T., Terada, M., & Sasaki, H. (2003). A faithful method for PCR-mediated global mRNA amplification and its integration into microarray analysis on laser-captured cells. Biochemical and Biophysical Research Communications, 300(4), 915-920. https://doi.org/10.1016/S0006-291X(02)02967-4