Title |
In Vitro Assembly of Multiple DNA Fragments Using Successive Hybridization
|
---|---|
Published in |
PLOS ONE, January 2012
|
DOI | 10.1371/journal.pone.0030267 |
Pubmed ID | |
Authors |
Xinglin Jiang, Jianming Yang, Haibo Zhang, Huibin Zou, Cong Wang, Mo Xian |
Abstract |
Construction of recombinant DNA from multiple fragments is widely required in molecular biology, especially for synthetic biology purposes. Here we describe a new method, successive hybridization assembling (SHA) which can rapidly do this in a single reaction in vitro. In SHA, DNA fragments are prepared to overlap one after another, so after simple denaturation-renaturation treatment they hybridize in a successive manner and thereby assemble into a recombinant molecule. In contrast to traditional methods, SHA eliminates the need for restriction enzymes, DNA ligases and recombinases, and is sequence-independent. We first demonstrated its feasibility by constructing plasmids from 4, 6 and 8 fragments with high efficiencies, and then applied it to constructing a customized vector and two artificial pathways. As SHA is robust, easy to use and can tolerate repeat sequences, we expect it to be a powerful tool in synthetic biology. |
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