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A recombinase-based palindrome generator capable of producing randomized shRNA libraries

Paper ID Volume ID Publish Year Pages File Format Full-Text
24593 43529 2009 6 PDF Available
Title
A recombinase-based palindrome generator capable of producing randomized shRNA libraries
Abstract

Short hairpin RNA (shRNA)-expressing vectors have been shown stably to knock-down directed targets through RNA interference (RNAi). RNAi has emerged as a powerful tool for reverse genetic screens by assaying cellular phenotypes after exposure to libraries of pooled shRNAs directed against all or a subset of cellular mRNAs. Recently, noncoding RNAs have been recognized as major arbiters of cellular phenotypes. The scope and diversity of noncoding RNAs greatly enlarge the target pool for reversible genetic screens and underscore the desirability as screening tools of complex RNAi libraries, such as randomized RNAi libraries. We describe a novel approach to generate randomized shRNAs that takes advantage of a stable plasmid intermediate that arises in a FLP recombinase system. The ability of this system to generate randomized palindromes represents a new technology for shRNA generation including random shRNAs that cannot be produced synthetically. We describe this plasmid system and its use in generating randomized shRNAs from input 20-bp oligomers, and validate the functionality of a shRNA produced using this approach to knock-down estrogen receptor alpha expression.

Keywords
FLP recombinase; Noncoding RNA; Randomized libraries; Interfering RNA
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A recombinase-based palindrome generator capable of producing randomized shRNA libraries
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 143, Issue 2, 20 August 2009, Pages 79–84
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us