期刊
ACS SYNTHETIC BIOLOGY
卷 6, 期 4, 页码 678-685出版社
AMER CHEMICAL SOC
DOI: 10.1021/acssynbio.6b00293
关键词
TALEN; DNA assembly; genome engineering; genetic screening; biofoundry; automation
资金
- Roy J. Carver Charitable Trust
- Carl R. Woese Institute for Genomic Biology at the University of Illinois at Urbana-Champaign (UIUC)
- Defense Advanced Research Projects Agency
- National Institutes of Health [1U54DK107965]
- 3M Corporation
- Carl R Woese Institute for Genomic Biology (UIUC)
Transcription activator-like effector nuclease (TALEN) is a programmable genome editing tool with wide applications. Since TALENs perform cleavage of DNA as heterodimers, a pair of TALENs must be synthesized for each target genome locus. Conventionally, TALEN pairs are either expressed on separate vectors or synthesized separately and then subcloned to the same vector. Neither approach allows high-throughput construction of TALEN libraries for largescale applications. Here we present a single-step assembly scheme to synthesize and express a pair of TALENs in a single transcript format with the help of a P2A self-cleavage sequence. Furthermore, we developed a fully automated platform to custom manufacture TALENs in a versatile biological foundry. 400 pairs of TALENs can be synthesized with over 96.2% success rate at a material cost of $2.1/pair. This platform opens the door to TALEN-based genome-wide studies.
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