High-Throughput Plasmid cDNA Library Screening
Libraries of cDNA clones are valuable resources foranalysing the expression, structure, and regulation of genes, as well asfor studying protein functions and interactions. Full-length cDNA clonesprovide information about intron and exon structures, splice junctionsand 5'- and 3'-untranslated regions (UTRs). Open reading frames (ORFs)derived from cDNA clones can be used to generate constructs allowingexpression of native proteins and N- or C-terminally tagged proteins.Thus, obtaining full-length cDNA clones and sequences for most or allgenes in an organism is critical for understanding genome functions.Expressed sequence tag (EST) sequencing samples cDNA libraries at random,which is most useful at the beginning of large-scale screening projects.However, as projects progress towards completion, the probability ofidentifying unique cDNAs via EST sequencing diminishes, resulting in poorrecovery of rare transcripts. We describe an adapted, high-throughputprotocol intended for recovery of specific, full-length clones fromplasmid cDNA libraries in five days.
- Research Organization:
- Ernest Orlando Lawrence Berkeley NationalLaboratory, Berkeley, CA (US)
- Sponsoring Organization:
- USDOE Director, Office of Science; National Institutes ofHealth
- DOE Contract Number:
- AC02-05CH11231
- OSTI ID:
- 923335
- Report Number(s):
- LBNL--60261; BnR: 400412000
- Journal Information:
- Nature Protocols, Journal Name: Nature Protocols Journal Issue: 0 Vol. 0
- Country of Publication:
- United States
- Language:
- English
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