Construction of recombinant DNA by exonuclease recession.

Abstract:

:We describe a new exonuclease-based method for joining and/or constructing two or more DNA molecules. DNA fragments containing ends complementary to those of a vector or another independent molecules were generated by the polymerase chain reaction. The 3' ends of these molecules as well as the vector DNA were then recessed by exonuclease activity and annealed in an orientation-determined manner via their complementary single-stranded regions. This recombinant DNA can be transformed directly into bacteria without a further ligase-dependent reaction. Using this approach, we have constructed recombinant DNA molecules rapidly, efficiently and directionally. This method can effectively replace conventional protocols for PCR cloning, PCR SOEing, DNA subcloning and site-directed mutagenesis.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Yang YS,Watson WJ,Tucker PW,Capra JD

doi

10.1093/nar/21.8.1889

subject

Has Abstract

pub_date

1993-04-25 00:00:00

pages

1889-93

issue

8

eissn

0305-1048

issn

1362-4962

journal_volume

21

pub_type

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