Subtraction of cap-trapped full-length cDNA libraries to select rare transcripts

Tomoko Hirozane-Kishikawa, Toshiyuki Shiraki, Kazunori Waki, Mari Nakamura, Takahiro Arakawa, Jun Kawai, Michela Fagiolini, Takao K. Hensch, Yoshihide Hayashizaki, Piero Carninci

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

The normalization and subtraction of highly expressed cDNAs from relatively large tissues before cloning dramatically enhanced the gene discovery by sequencing for the mouse full-length cDNA encyclopedia, but these methods have not been suitable for limited RNA materials. To normalize and subtract full-length cDNA libraries derived from limited quantities of total RNA, here we report a method to subtract plasmid libraries excised from size-unbiased amplified λ phage cDNA libraries that avoids heavily biasing steps such as PCR and plasmid library amplification. The proportion of full-length cDNAs and the gene discovery rate are high, and library diversity can be validated by in silico randomization.

Original languageEnglish
Pages (from-to)510-518
Number of pages9
JournalBioTechniques
Volume35
Issue number3
DOIs
Publication statusPublished - 2003 Sep 1

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)

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