A One-Step System for Convenient and Flexible Assembly of Transcription Activator-Like Effector Nucleases (TALENs)

Zhao Wu; Jinlong Zhao; Wenye Sun; Jing Liang; Jing Jiang

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자료유형: 학술저널

저자정보: Zhao Wu (From Innovative Cellular Therapeutics Co) Jinlong Zhao (From Innovative Cellular Therapeutics Co) Wenye Sun (From Innovative Cellular Therapeutics Co) Jing Liang (From Innovative Cellular Therapeutics Co) Jing Jiang (From Innovative Cellular Therapeutics Co)

저널정보: 한국분자세포생물학회 Molecules and Cells Molecules and Cells 제39권 제9호

발행연도: 2016.9

수록면: 687 - 691 (5page)

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Transcription activator-like effector nucleases (TALENs) are powerful tools for targeted genome editing in diverse cell types and organisms. However, the highly identical TALE repeat sequences make it challenging to assemble TALEs using conventional cloning approaches, and multiple repeats in one plasmid are easily catalyzed for homologous recombination in bacteria. Although the methods for TALE assembly are constantly improving, these methods are not convenient because of laborious assembly steps or large module libraries, limiting their broad utility. To overcome the barrier of multiple assembly steps, we report a one-step system for the con-venient and flexible assembly of a 180 TALE module library. This study is the first demonstration to ligate 9 mono-/dimer modules and one circular TALEN backbone vector in a one step process, generating 9.5 to 18.5 repeat sequences with an overall assembly rate higher than 50%. This system makes TALEN assembly much simpler than the conventional cloning of two DNA fragments because this strategy combines digestion and ligation into one step using circular vectors and different modules to avoid gel extraction. Therefore, this system provides a convenient tool for the application of TALEN-mediated genome editing in scientific studies and clinical trials.

#genome editing #molecular genetics #transcription activator-like effector nuclease (TALEN)

참고문헌 (14)

참고문헌 신청

Boch, J. / 2009 / Breaking the code of DNA binding specificity of TAL-type III effectors / Science 326:1509~1512

Cermak, T. / 2011 / Efficient design and assembly of custom TALEN and other TAL effector-based constructs for DNA targeting / Nucleic Acids Res 39:82

Ding, Q. / 2013 / A TALEN genome-editing system for generating human stem cell-based disease models / Cell Stem Cell 12:238~251

Engler, C. / 2008 / A one pot, one step, precision cloning method with high throughput capability / PLoS One 3:3647

Engler, C. / 2009 / Golden gate shuffling : a one-pot DNA shuffling method based on type IIs restriction enzymes / PLoS One 4:5553

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