High-throughput fluorescence labelling of full-length cDNA products based on a reconstituted translation system

Yuko Kawahashi, Nobuhide Doi, Yo Oishi, Chizuru Tsuda, Hideaki Takashima, Tomoya Baba, Hirotada Mori, Takashi Ito, Hiroshi Yanagawa

研究成果: Article査読

4 被引用数 (Scopus)

抄録

Although recent advances in fluorescence-based technologies, such as protein microarrays, have made it possible to analyse more than 10,000 proteins at once, there is a bottleneck in the step of preparation of large numbers of fluorescently labelled proteins for the comprehensive analysis of protein-protein interactions. Here we describe two independent methods for high-throughput fluorescence-labelling of full-length cDNA products at their C-termini using a reconstituted translation system containing fluorescent puromycin. For the first method, release factor-free systems were used. For the second method, stop codons were excluded from cDNAs by using a common mismatch primer in mutagenic PCR. These methods yielded large numbers of labelled proteins from cDNA sets of various organisms, such as mouse, yeast and Escherichia coli.

本文言語English
ページ(範囲)19-24
ページ数6
ジャーナルJournal of biochemistry
141
1
DOI
出版ステータスPublished - 2007 1

ASJC Scopus subject areas

  • 生化学
  • 分子生物学

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