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dc.contributor.authorBriggs, Adrian W.
dc.contributor.authorRios, Xavier
dc.contributor.authorChari, Raj
dc.contributor.authorYang, Luhan
dc.contributor.authorZhang, Feng
dc.contributor.authorMali, Prashant
dc.contributor.authorChurch, George M.
dc.date.accessioned2016-05-25T18:04:58Z
dc.date.available2016-05-25T18:04:58Z
dc.date.issued2012-06
dc.date.submitted2012-06
dc.identifier.issn0305-1048
dc.identifier.issn1362-4962
dc.identifier.urihttp://hdl.handle.net/1721.1/102681
dc.description.abstractDNA built from modular repeats presents a challenge for gene synthesis. We present a solid surface-based sequential ligation approach, which we refer to as iterative capped assembly (ICA), that adds DNA repeat monomers individually to a growing chain while using hairpin ‘capping’ oligonucleotides to block incompletely extended chains, greatly increasing the frequency of full-length final products. Applying ICA to a model problem, construction of custom transcription activator-like effector nucleases (TALENs) for genome engineering, we demonstrate efficient synthesis of TALE DNA-binding domains up to 21 monomers long and their ligation into a nuclease-carrying backbone vector all within 3 h. We used ICA to synthesize 20 TALENs of varying DNA target site length and tested their ability to stimulate gene editing by a donor oligonucleotide in human cells. All the TALENS show activity, with the ones >15 monomers long tending to work best. Since ICA builds full-length constructs from individual monomers rather than large exhaustive libraries of pre-fabricated oligomers, it will be trivial to incorporate future modified TALE monomers with improved or expanded function or to synthesize other types of repeat-modular DNA where the diversity of possible monomers makes exhaustive oligomer libraries impractical.en_US
dc.description.sponsorshipNational Human Genome Research Institute (U.S.) ((NHGRI) 1P50 HG005550)en_US
dc.description.sponsorshipEuropean Molecular Biology Organization (ALTF 91-2010)en_US
dc.description.sponsorshipCanadian Institutes of Health Research (Banting Postdoctoral fellowship)en_US
dc.language.isoen_US
dc.publisherOxford University Pressen_US
dc.relation.isversionofhttp://dx.doi.org/10.1093/nar/gks624en_US
dc.rightsCreative Commons Attribution Non-Commercial Licenseen_US
dc.rights.urihttp://creativecommons.org/licenses/by-nc/3.0/en_US
dc.sourceOxford University Pressen_US
dc.titleIterative capped assembly: rapid and scalable synthesis of repeat-module DNA such as TAL effectors from individual monomersen_US
dc.typeArticleen_US
dc.identifier.citationBriggs, A. W., X. Rios, R. Chari, L. Yang, F. Zhang, P. Mali, and G. M. Church. “Iterative Capped Assembly: Rapid and Scalable Synthesis of Repeat-Module DNA Such as TAL Effectors from Individual Monomers.” Nucleic Acids Research 40, no. 15 (June 26, 2012): e117–e117.en_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Brain and Cognitive Sciencesen_US
dc.contributor.mitauthorZhang, Fengen_US
dc.relation.journalNucleic Acids Researchen_US
dc.eprint.versionFinal published versionen_US
dc.type.urihttp://purl.org/eprint/type/JournalArticleen_US
eprint.statushttp://purl.org/eprint/status/PeerRevieweden_US
dspace.orderedauthorsBriggs, A. W.; Rios, X.; Chari, R.; Yang, L.; Zhang, F.; Mali, P.; Church, G. M.en_US
dspace.embargo.termsNen_US
dc.identifier.orcidhttps://orcid.org/0000-0003-2782-2509
mit.licensePUBLISHER_CCen_US


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