Abstract:
:The ability to rapidly design, build, and test prototypes is of key importance to every engineering discipline. DNA assembly often serves as a rate limiting step of the prototyping cycle for synthetic biology. Recently developed DNA assembly methods such as isothermal assembly and type IIS restriction enzyme systems take different approaches to accelerate DNA construction. We introduce a hybrid method, Golden Gate-Gibson (3G), that takes advantage of modular part libraries introduced by type IIS restriction enzyme systems and isothermal assembly's ability to build large DNA constructs in single pot reactions. Our method is highly efficient and rapid, facilitating construction of entire multigene circuits in a single day. Additionally, 3G allows generation of variant libraries enabling efficient screening of different possible circuit constructions. We characterize the efficiency and accuracy of 3G assembly for various construct sizes, and demonstrate 3G by characterizing variants of an inducible cell-lysis circuit.
journal_name
ACS Synth Bioljournal_title
ACS synthetic biologyauthors
Halleran AD,Swaminathan A,Murray RMdoi
10.1021/acssynbio.8b00060subject
Has Abstractpub_date
2018-05-18 00:00:00pages
1477-1480issue
5issn
2161-5063journal_volume
7pub_type
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