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Nat Genet. 2004 Feb;36(2):190-6. Epub 2004 Jan 4.

Enzymatic production of RNAi libraries from cDNAs.

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Department of Pharmacology, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.


RNA interference (RNAi) induced by small interfering (siRNA) or short hairpin RNA (shRNA) is an important research approach in mammalian genetics. Here we describe a technology called enzymatic production of RNAi library (EPRIL) by which cDNAs are converted by a sequence of enzymatic treatments into an RNAi library consisting of a vast array of different shRNA expression constructs. We applied EPRIL to a single cDNA source and prepared an RNAi library consisting of shRNA constructs with various RNAi efficiencies. High-throughput screening allowed us to rapidly identify the best shRNA constructs from the library. We also describe a new selection scheme using the thymidine kinase gene for obtaining efficient shRNA constructs. Furthermore, we show that EPRIL can be applied to constructing an RNAi library from a cDNA library, providing a basis for future whole-genome phenotypic screening of genes.

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