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  A simple TALEN-based protocol for efficient genome-editing in Drosophila

Zhang, X., Ferreira, I. R. S., & Schnorrer, F. (2014). A simple TALEN-based protocol for efficient genome-editing in Drosophila. METHODS, 69(1), 32-37. doi:10.1016/j.ymeth.2014.03.020.

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 Creators:
Zhang, Xu1, Author              
Ferreira, Irene R. S.1, Author              
Schnorrer, Frank1, Author              
Affiliations:
1Schnorrer, Frank / Muscle Dynamics, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565168              

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Free keywords: ZINC-FINGER NUCLEASES; HOMOLOGOUS RECOMBINATION; TRANSGENIC RNAI; MELANOGASTER; MUTATIONS; DESIGNDrosophila; Mutagenesis; Genome-editing; TALEN; T7 endonuclease I; Method;
 Abstract: Drosophila is a well-established genetic model organism: thousands of point mutations, deficiencies or transposon insertions are available from stock centres. However, to date, it is still difficult to modify a specific gene locus in a defined manner. A potential solution is the application of transcription activator-like effector nucleases (TALENs), which have been used successfully to mutate genes in various model organisms. TALENs are constructed by fusion of TALE proteins to the endonuclease FokI, resulting in artificial, sequence-specific endonucleases. They induce double strand breaks, which are either repaired by error-prone non-homologous end joining (NHEJ) or homology directed repair (HDR). We developed a simple TALEN-based protocol to mutate any gene of interest in Drosophila within approximately 2 months. We inject mRNA coding for two TALEN pairs targeting the same gene into embryos, employ T7 endonuclease I screening of pooled F-1 flies to identify mutations and generate a stable mutant stock in the F-3 generation. We illustrate the efficacy of our strategy by mutating CG11617, a previously uncharacterized putative transcription factor with an unknown function in Drosophila. This demonstrates that TALENs are a reliable and efficient strategy to mutate any gene of interest in Drosophila. (C) 2014 Elsevier Inc. All rights reserved.

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Language(s): eng - English
 Dates: 2014
 Publication Status: Published in print
 Pages: 6
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Degree: -

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Title: METHODS
Source Genre: Journal
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Publ. Info: 525 B ST, STE 1900, SAN DIEGO, CA 92101-4495 USA : ACADEMIC PRESS INC ELSEVIER SCIENCE
Pages: - Volume / Issue: 69 (1) Sequence Number: - Start / End Page: 32 - 37 Identifier: ISSN: 1046-2023