Large-scale concatenation cDNA sequencing

• Published in: Genome research 1997-04, Vol.7 (4), p.353-358
• Main Authors: Yu, W, Andersson, B, Worley, K C, Muzny, D M, Ding, Y, Liu, W, Ricafrente, J Y, Wentland, M A, Lennon, G, Gibbs, R A
• Format: Article
• Language: English
• Subjects: Databases, Factual; DNA Transposable Elements; DNA, Complementary - chemistry; DNA, Complementary - genetics; Gene Library; Humans; Letters; Molecular Sequence Data; Proteins - chemistry; Proteins - genetics; Sequence Alignment - methods; Sequence Analysis, DNA - methods; Sequence Homology, Amino Acid; Sequence Homology, Nucleic Acid; Software
• ISSN: 1088-9051; 1549-5469
• DOI: 10.1101/gr.7.4.353

A total of 100 kb of DNA derived from 69 individual human brain cDNA clones of 0.7-2.0 kb were sequenced by concatenated cDNA sequencing (CCS), whereby multiple individual DNA fragments are sequenced simultaneously in a single shotgun library. The method yielded accurate sequences and a similar efficiency compared with other shotgun libraries constructed from single DNA fragments (> 20 kb). Computer analyses were carried out on 65 cDNA clone sequences and their corresponding end sequences to examine both nucleic acid and amino acid sequence similarities in the databases. Thirty-seven clones revealed no DNA database matches, 12 clones generated exact matches (> or = 98% identity), and 16 clones generated nonexact matches (57%-97% identity) to either known human or other species genes. Of those 28 matched clones, 8 had corresponding end sequences that failed to identify similarities. In a protein similarity search, 27 clone sequences displayed significant matches, whereas only 20 of the end sequences had matches to known protein sequences. Our data indicate that full-length cDNA insert sequences provide significantly more nucleic acid and protein sequence similarity matches than expressed sequence tags (ESTs) for database searching.