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Genome Res. 2000 Oct;10(10):1617-30.

Normalization and subtraction of cap-trapper-selected cDNAs to prepare full-length cDNA libraries for rapid discovery of new genes.

Author information

1
Laboratory for Genome Exploration Research Group, RIKEN Genomic Sciences Center, Tsukuba, Japan. rgscerg@rtc.riken.go.jp

Abstract

In the effort to prepare the mouse full-length cDNA encyclopedia, we previously developed several techniques to prepare and select full-length cDNAs. To increase the number of different cDNAs, we introduce here a strategy to prepare normalized and subtracted cDNA libraries in a single step. The method is based on hybridization of the first-strand, full-length cDNA with several RNA drivers, including starting mRNA as the normalizing driver and run-off transcripts from minilibraries containing highly expressed genes, rearrayed clones, and previously sequenced cDNAs as subtracting drivers. Our method keeps the proportion of full-length cDNAs in the subtracted/normalized library high. Moreover, our method dramatically enhances the discovery of new genes as compared to results obtained by using standard, full-length cDNA libraries. This procedure can be extended to the preparation of full-length cDNA encyclopedias from other organisms.

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PMID:
11042159
PMCID:
PMC310980
DOI:
10.1101/gr.145100
[Indexed for MEDLINE]
Free PMC Article

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