pGP-B2E, a Recombinant Compatible TA/TB-Ligation Vector for Rapid and Inexpensive Gene Cloning
- 15 Downloads
DNA cloning is the basic step for different fields of life science, and many efforts have been made to simplify this procedure. In this study, we report two general purpose plasmids (pGP), pGP-XB2E and pGP-B2E, for rapid and cost-effective construct of basic clones. The BciVI and XcmI cleavage sites are designed in pGP-XB2E to test plasmid linearization efficiency. The plasmid has better linearization efficiency by using BciVI which could almost completely digest 2 μg plasmid in 10 min with only one-tenth the recommended enzyme concentration. In order to further optimize the pGP-XB2E, a new plasmid, pGP-B2E, which removed XcmI cleavage site was designed. This vector is highly efficient for cloning PCR products up to 1812 bp, and the number of colonies was about five times that of pGP-XB2E. In addition to TA cloning, blunt-end PCR products with T ended in the primer could be positively linked to the T-vector pGP-B2E without A-tailing treatment (TB cloning). Moreover, as an entry vector, pGP-B2E was also compatible for gateway recombination reaction without frameshift mutations. In general, this vector provides a universal, quick, and cost-efficient method for basic molecular cloning.
KeywordsTA cloning Blunt-end ligation Gateway entry vector High efficiency Low cost
This work was supported by the National Key Research and Development Program of China (2016YFD0100601-15), Zhejiang Provincial Foundation for Natural Science (LZ16C130002, LY16C140005, LY17C130007), and Zhejiang Fundamental Public Welfare Research Program (LGN19C140008).
Compliance with Ethical Standards
Conflict of interest
The authors declare no competing or financial interest.
- 26.Zhou, J., Wang, X. M., Chen, B., Chen, J., Yang, Y., Yu, C. L., et al. (2013). Low-cost gateway-compatible bimolecular fluorescence complementation assay system. Acta Agriculturae Zhejiangensis,25, 1024–1030.Google Scholar