Abstract：
:The recently evolved field of synthetic biology has revolutionized the way we think of biology as an "engineerable" discipline. The newly sprouted branch is constantly in need of simple, cost-effective and automatable DNA-assembly methods. We have developed a reliable DNA-assembly system, ZeBRα (Zero-Background Redα), for cloning multiple DNA-fragments seamlessly with very high efficiency. The hallmarks of ZeBRα are the greatly reduced hands-on time and costs and yet excellent efficiency and flexibility. ZeBRα combines a "zero-background vector" with a highly efficient in vitro recombination method. The suicide-gene in the vector acts as placeholder, and is replaced by the fragments-of-interest, ensuring the exclusive survival of the successful recombinants. Thereby the background from uncut or re-ligated vector is absent and screening for recombinant colonies is unnecessary. Multiple fragments-of-interest can be assembled into the empty vector by a recombinogenic E. coli-lysate (SLiCE) with a total time requirement of less than 48 h. We have significantly simplified the preparation of the high recombination-competent E. coli-lysate compared to the original protocol. ZeBRα is the least labor intensive among comparable state-of-the-art assembly/cloning methods without a trade-off in efficiency.
journal_name
Sci Repjournal_title
Scientific reportsauthors
Richter D,Bayer K,Toesko T,Schuster Sdoi
10.1038/s41598-019-39768-0subject
Has Abstractpub_date
2019-02-27 00:00:00pages
2980issue
1issn
2045-2322pii
10.1038/s41598-019-39768-0journal_volume
9pub_type
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