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A Framework for piRNA Cluster Manipulation

Olovnikov, Ivan and Le Thomas, Adrien and Aravin, Alexei A. (2014) A Framework for piRNA Cluster Manipulation. In: PIWI-Interacting RNAs: Methods and Protocols. Methods in Molecular Biology. No.1093. Humana Press , New York, NY, pp. 47-58. ISBN 978-1-62703-693-1.

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Piwi proteins and their small-RNA partners, piwi-interacting (pi)RNA, form a natural mechanism that prevents the deleterious activity of transposable elements in the germ line of metazoan species. The piRNA pathway relies on extended noncoding genomic regions, dubbed piRNA clusters, to produce long precursor transcripts that are subsequently processed into mature piRNAs. The large size and repetitive nature of piRNA clusters provide significant challenges for their dissection using common genetic tools. Here we describe an effective approach for manipulation of piRNA clusters using a combination of BAC recombineering in E. coli and phiC31-mediated transgenesis in Drosophila. Although the described approach is instrumental for manipulating piRNA clusters, it can also be implemented for other problems in functional genomics.

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Additional Information:© 2014 Springer Science+Business Media, LLC. We thank members of the Aravin lab for helpful discussion and comments on the manuscript. We are particularly thankful to Alexandre Webster for comments and editing. I.O. is a CEMI (Center for Environmental Microbiology Interactions) fellow at Caltech. This work was supported by grants from the National Institutes of Health (R01 GM097363, R00 HD057233, and DP2 OD007371A) and the Searle Scholar Award to A.A.A.
Group:Caltech Center for Environmental Microbial Interactions (CEMI)
Funding AgencyGrant Number
NIHR01 GM097363
NIHR00 HD057233
NIHDP2 OD007371A
Searle Scholar AwardUNSPECIFIED
Caltech Center for Environmental Microbial Interactions (CEMI)UNSPECIFIED
Subject Keywords:piRNA, piRNA cluster, BAC, Recombineering, phiC31
Series Name:Methods in Molecular Biology
Issue or Number:1093
Record Number:CaltechAUTHORS:20131112-093205322
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:42373
Deposited By: Ruth Sustaita
Deposited On:12 Nov 2013 17:59
Last Modified:10 Nov 2021 16:22

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