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High-efficiency cloning of Arabidopsis full-length cDNA by biotinylated CAP trapper

Authors

  • Motoaki Seki,

    1. Laboratory of Plant Molecular Biology, Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), 3–1–1 Koyadai, Tsukuba 305–0074, Japan,
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  • Piero Carninci,

    1. Genome Science Laboratory, Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), 3–1–1 Koyadai, Tsukuba 305–0074, Japan, and
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  • Yoko Nishiyama,

    1. Genome Science Laboratory, Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), 3–1–1 Koyadai, Tsukuba 305–0074, Japan, and
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  • Yoshihide Hayashizaki,

    1. Genome Science Laboratory, Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), 3–1–1 Koyadai, Tsukuba 305–0074, Japan, and
    2. CREST, Japan Science and Technology Corporation (JST), Kawaguchi-Center building, 4–1–8 Hon-cho, Kawaguchi, Saitama 332, Japan
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  • Kazuo Shinozaki

    1. Laboratory of Plant Molecular Biology, Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), 3–1–1 Koyadai, Tsukuba 305–0074, Japan,
    2. CREST, Japan Science and Technology Corporation (JST), Kawaguchi-Center building, 4–1–8 Hon-cho, Kawaguchi, Saitama 332, Japan
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*For correspondence (fax +81 29836 9060; e-mail sinozaki@rtc.riken.go.jp).

Summary

Full-length cDNAs are essential for functional analysis of plant genes. We constructed high-content, full-length cDNA libraries from Arabidopsis thaliana plants based on chemical introduction of a biotin group into the diol residue of the CAP structure of eukaryotic mRNA, followed by RNase I treatment, to select full-length cDNA. More than 90% of the total clones obtained were of full length; recombinant clones were obtained with high efficiency (2.2 × 106/9 μg starting mRNA). Sequence analysis of 111 randomly picked clones indicated that 32 isolated cDNA groups were derived from novel genes in the A. thaliana genome.

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