title: Fully Automated One-Step Synthesis of Single-Transcript TALEN Pairs Using a Biological Foundry.
abstract: 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 large-scale 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.
authors.affiliation: Carl R. Woese Institute for Genomic Biology, ‡Department of Chemical and Biomolecular Engineering, §Department of Biochemistry, ∥Department of Microbiology, ⊥Department of Chemistry, and #Department of Bioengineering, University of Illinois at Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Ran
authors.initials: R
authors.lastname: Chao
authors.affiliation: Carl R. Woese Institute for Genomic Biology, ‡Department of Chemical and Biomolecular Engineering, §Department of Biochemistry, ∥Department of Microbiology, ⊥Department of Chemistry, and #Department of Bioengineering, University of Illinois at Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Jing
authors.initials: J
authors.lastname: Liang
authors.affiliation: Carl R. Woese Institute for Genomic Biology, ‡Department of Chemical and Biomolecular Engineering, §Department of Biochemistry, ∥Department of Microbiology, ⊥Department of Chemistry, and #Department of Bioengineering, University of Illinois at Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Ipek
authors.initials: I
authors.lastname: Tasan
authors.affiliation: Carl R. Woese Institute for Genomic Biology, ‡Department of Chemical and Biomolecular Engineering, §Department of Biochemistry, ∥Department of Microbiology, ⊥Department of Chemistry, and #Department of Bioengineering, University of Illinois at Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Tong
authors.identifier: 0000-0003-2985-9014
authors.initials: T
authors.lastname: Si
authors.affiliation: Carl R. Woese Institute for Genomic Biology, ‡Department of Chemical and Biomolecular Engineering, §Department of Biochemistry, ∥Department of Microbiology, ⊥Department of Chemistry, and #Department of Bioengineering, University of Illinois at Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Linyang
authors.initials: L
authors.lastname: Ju
authors.affiliation: Carl R. Woese Institute for Genomic Biology, ‡Department of Chemical and Biomolecular Engineering, §Department of Biochemistry, ∥Department of Microbiology, ⊥Department of Chemistry, and #Department of Bioengineering, University of Illinois at Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Huimin
authors.identifier: 0000-0002-9069-6739
authors.initials: H
authors.lastname: Zhao
mesh_terms: D001331:Automation; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D000072669:Gene Editing / Q000295:instrumentation / Q000379:methods*; D055786:Gene Knockout Techniques; D057809:HEK293 Cells; D006801:Humans; D000069896:Transcription Activator-Like Effector Nucleases / Q000235:genetics / Q000378:metabolism*
chemical_list: D004247:DNA; D000069896:Transcription Activator-Like Effector Nucleases
keywords: DNA assembly; TALEN; automation; biofoundry; genetic screening; genome engineering