title: Full-length transcriptome analysis of human retina-derived cell lines ARPE-19 and Y79 using the vector-capping method.
abstract: PURPOSE. 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. METHODS. 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. RESULTS. 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. CONCLUSIONS. 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.
authors.affiliation: Department of Rehabilitation Engineering, Research Institute, National Rehabilitation Center for Persons with Disabilities, Tokorozawa, Japan.
authors.forename: Mio
authors.initials: M
authors.lastname: Oshikawa
authors.forename: Chihiro
authors.initials: C
authors.lastname: Tsutsui
authors.forename: Tomoko
authors.initials: T
authors.lastname: Ikegami
authors.forename: Yuki
authors.initials: Y
authors.lastname: Fuchida
authors.forename: Maki
authors.initials: M
authors.lastname: Matsubara
authors.forename: Shigeru
authors.initials: S
authors.lastname: Toyama
authors.forename: Ron
authors.initials: R
authors.lastname: Usami
authors.forename: Kuniyo
authors.initials: K
authors.lastname: Ohtoko
authors.forename: Seishi
authors.initials: S
authors.lastname: Kato
mesh_terms: D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D015870:Gene Expression / Q000502:physiology*; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D015723:Gene Library; D052138:Genes, Neoplasm; D005822:Genetic Vectors; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D010957:Plasmids; D012315:RNA Caps / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D019572:Retinal Neoplasms / Q000235:genetics*; D055213:Retinal Pigment Epithelium / Q000378:metabolism*; D012175:Retinoblastoma / Q000235:genetics*; D014158:Transcription, Genetic
chemical_list: D012315:RNA Caps; D012333:RNA, Messenger