Enzyme Treatment-Free and Ligation-Independent Cloning Using Caged Primers in Polymerase Chain Reactions

Molecules

Akinori Kuzuya¹  Keita Tanaka¹  Hitoshi Katada¹ 

[1]¹Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904, Japan[2] height:6px

[3]"> ²Department of Chemistry and Materials Engineering, Kansai University, 3-3-35 Yamate-cho, Suita, Osaka 564-8680, Japan

关键词: DNA;    PCR;    caged compounds;    ligation;    cloning;   

DOI  :  10.3390/molecules17010328

来源: mdpi

【 摘 要 】

A new simple scheme for constructing recombinant vectors that does not require any restriction enzyme, ligase, or any other special enzyme treatment has been developed. By using caged primers in PCR, unnatural sticky-ends of any sequence, which are sufficiently long for ligation-independent cloning (LIC), are directly prepared on the product after a brief UVA irradiation. Target genes and vectors amplified by this light-assisted cohesive-ending (LACE) PCR join together in the desired arrangement in a simple mixture of them, tightly enough to be repaired and ligated in competent cells.

【 授权许可】

CC BY   
This is an open access article distributed under the Creative Commons Attribution License (CC BY) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.