Simultaneous shotgun sequencing of multiple cDNA clones.

TitleSimultaneous shotgun sequencing of multiple cDNA clones.
Publication TypeJournal Article
Year of Publication1997
AuthorsAndersson, B, Lu, J, Shen, Y, Wentland, MA, Gibbs, RA
JournalDNA Seq
Volume7
Issue2
Pagination63-70
Date Published1997
ISSN1042-5179
KeywordsAnimals, Cloning, Molecular, DNA, Complementary, Drosophila, Gene Expression, Gene Library, Restriction Mapping, Sequence Analysis, DNA
Abstract

This study describes an efficient method for the rapid sequencing of DNA fragments in the size range 1-5 kb. Individual fragments, here cDNA inserts, are purified by restriction enzyme digestion and gel purification, pooled and concatenated by ligation. The concatamers are sheared and cloned randomly into M13, followed by random sequencing. The sequences of the individual cDNA inserts are obtained at the assembly stage using restriction enzyme sites as "tags' for the ends of each fragment. In this study the sequencing of two libraries containing 7 and 16 cDNA inserts with an average length of 2.5 kb is described. The results show that of the shotgun sequencing of large fragments, and that the method compares favorably to alternative strategies, such as primer walking.

Alternate JournalDNA Seq.
PubMed ID9063643
Grant ListP30 HG00210 / HG / NHGRI NIH HHS / United States
R01 HG00823 / HG / NHGRI NIH HHS / United States