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Mio Oshikawa, Chihiro Tsutsui, Tomoko Ikegami, Yuki Fuchida, Maki Matsubara, Shigeru Toyama, Ron Usami, Kuniyo Ohtoko, Seishi Kato; Full-Length Transcriptome Analysis of Human Retina-Derived Cell Lines ARPE-19 and Y79 Using the Vector-Capping Method. Invest. Ophthalmol. Vis. Sci. 2011;52(9):6662-6670. doi: 10.1167/iovs.11-7479.
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© ARVO (1962-2015); The Authors (2016-present)
To collect an entire set of full-length cDNA clones derived from human retina-derived cell lines and to identify full-length transcripts for retinal preferentially expressed genes.
The full-length cDNA libraries were constructed from a retinoblastoma cell line, Y79, and a retinal pigment epithelium cell line, ARPE-19, using the vector-capping method, which generates a genuine full-length cDNA. By single-pass sequencing of the 5′-end of cDNA clones and subsequent mapping to the human genome, the authors determined their transcriptional start sites and annotated the cDNA clones.
Of the 23,616 clones isolated from Y79-derived cDNA libraries, 19,229 full-length cDNA clones were identified and classified into 4808 genes, including genes of >10 kbp. Of the 7067 genes obtained from the Y79 and ARPE-19 libraries, the authors selected 72 genes that were preferentially expressed in the eye, of which 131 clones corresponding to 57 genes were fully sequenced. As a result, we discovered many variants that were produced by different transcriptional start sites, alternative splicing, and alternative polyadenylation.
The bias-free, full-length cDNA libraries constructed using the vector-capping method were shown to be useful for collecting an entire set of full-length cDNA clones for these retinal cell lines. Full-length transcriptome analysis of these cDNA libraries revealed that there were, unexpectedly, many transcript variants for each gene, indicating that obtaining the full-length cDNA for each variant is indispensable for analyzing its function. The full-length cDNA clones (approximately 80,000 clones each for ARPE-19 and Y79) will be useful as a resource for investigating the human retina.
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