Construction and characterization of a full-length cDNA library in Chlamydomonas reinhardtii

Takeaki Kubo1, Tomoya Satake1, Katsuyuki Yamato1, Takashi Yamano1, Yutaka Suzuki2, Sumio Sugano2, Takehiko Ito3, Tadasu Shin-i4, Yuji Kohara4, Asao Fujiyama5, and Hideya Fukuzawa1

1Graduate school of Biostudies, Kyoto Univ., Kyoto 606-8502, Japan 2International and Interdisciplinary Studies, Univ. of Tokyo, Japan 3Mitsubishi Research Institute, Tokyo, Japan 4National Institute of Informatics, Tokyo, Japan 5National Institute of Genetics, Mishima, Japan

By using the gene prediction algorithms and integrated with EST sequences generated from various environmental conditions, a candidate set of 15,143 protein coding gene was predicted on the draft genome sequence of C. reinhardtii (Merchant et al. 2007). However, in some cases the gene models lack 5'-ends of mRNA coding regions, or parts of exons. Therefore, it is important to collect the full-covered mRNA sequences of each gene to predict precise mRNA sequence leading to actual peptide sequences of the gene products. In order to collect the gene-set with polypeptide sequences on Chlamydomonas genome, a full length-cDNA enriched library was constructed by using oligo-capping method (Suzuki et al. 1997). The mRNAs for the library were isolated from various environment stresses such as high light, low temperature, high osmolality, limitation of micro and macro nutrients, deflagellation and sexual/asexual cycle. Both ends of the cDNA sequences, in total 107,970 reads, were assembled with preciously known mRNA sequences deposited in NCBI and ACEGs predicted from the JGI genome sequence (v.3.0) (Jain et al. 2007). As a result, 2,800 clusters covered by full length cDNAs were identified from the 14,124 predicted clusters. The analysis of the predicted peptide sequences encoded by the full-length cDNA is in progress.

e-mail address of presenting author: kubot@lif.kyoto-u.ac.jp