NDLI: Vectors and parameters that enhance the efficacy of RNAi-mediated gene disruption in transgenic Drosophila

Content Provider	PubMed Central
Author	Haley, Benjamin Foys, Bryon Levine, Michael
Abstract	Whole-genome transgenic RNAi libraries permit systematic genetic screens in individual tissues of Drosophila. However, there is a high incidence of nonspecific phenotypes because of off-target effects. To minimize such effects, it is essential to obtain a deeper understanding of the specificity of action of RNAi. Here, in vivo assays are used to determine the minimum, contiguous nucleotide pairing required between an siRNA and a target mRNA to generate a phenotype. We observe that as few as 16 nucleotides of contiguous homology are sufficient to attenuate gene activity. This finding provides an explanation for the high incidence of off-target effects observed in RNAi-based genetic screens. Toward improving the efficacy of RNAi-induced phenotypes in vivo, we describe siRNA expression vectors that allow coexpression of one or more siRNAs with a fluorescent reporter gene in cultured cells or transgenic flies. This expression system makes use of the small intron from the ftz segmentation gene to provide efficient processing of synthetic siRNAs from a reporter transcript. These studies provide a foundation for the specific and effective use of gene silencing in transgenic Drosophila.
Related Links	http://dx.doi.org/10.1073/pnas.1006689107
Ending Page	11440
Page Count	6
Starting Page	11435
File Format	PDF
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	25
Volume Number	107
Language	English
Publisher	National Academy of Sciences
Publisher Date	2010-06-22
Access Restriction	Open
Rights Holder	National Academy of Sciences
Subject Keyword	General Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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