Construction of two pGEM-7Zf(+) phagemid T-tail vectors using AhdI-restriction endonuclease sites for direct cloning of PCR products

For applications such as sequencing, transfection, and in vitro transcription, PCR products have to be subcloned into plasmids. Many strategies are used for cloning, blunt-end ligation or the incorporation of restriction endonuclease sites into PCR primers for appropriate vectors. However, the most...

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Bibliographic Details
Published in:Plasmid 2002-09, Vol.48 (2), p.160-163
Main Authors: Jeung, Ji Ung, Cho, Sung Ki, Shim, Kyu Suk, Ok, Sung Han, Lim, Dae Sik, Shin, Jeong Sheop
Format: Article
Language:English
Subjects:
Base Sequence
Cloning, Molecular - methods
DNA Restriction Enzymes - metabolism
Genetic Vectors - genetics
Polymerase Chain Reaction
Substrate Specificity
Transformation, Genetic
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Summary:For applications such as sequencing, transfection, and in vitro transcription, PCR products have to be subcloned into plasmids. Many strategies are used for cloning, blunt-end ligation or the incorporation of restriction endonuclease sites into PCR primers for appropriate vectors. However, the most convenient and direct method is T/A cloning. In this study, we developed two of the pGEM-7Zf(+) phagemid T-tail vectors using AhdI-restriction endonuclease sites, and these T vectors have all the features of pGEM-7Zf(+): f1 ori, T7, and SP6 RNA polymerase promoters, the alpha-peptide coding region of beta-galactosidase for X-gal blue/white color selection, the beta-lactamase gene for recombinant colony selection, and binding sites for pUC/M13 forward and reverse sequencing primers. These AhdI-containing phagemid vectors, pGEM-NJ105 and pGEM-NJ107, are useful for the easy and inexpensive preparation of T vectors and direct cloning of PCR products.
ISSN:0147-619X
1095-9890
DOI:10.1016/S0147-619X(02)00122-1