Promega, A3600, pGEM®-T Vector Systems

Efficient T-Vector Cloning with Blue/White Selection
The pGEM®-T Vector Systems are convenient systems to clone PCR products generated by certain thermostable polymerases. These polymerases often add a single deoxyadenosine, in a template-independent fashion, to the 3´-ends of the amplified fragments. The pGEM®-T Vector is ready to use in ligation reactions, prepared by cutting the pGEM®-5Zf(+) Vector with EcoRV and adding a 3´ terminal thymidine to both ends. These single 3´-T overhangs at the insertion site greatly improve the efficiency of ligation of a PCR product into the plasmid by preventing recircularization of the vector and providing a compatible overhang for ligation of PCR products with A overhangs.
pGEM®-T Vector Map and Sequence
The pGEM®-T Vector is derived from the pGEM®-5Zf(+) Vector (GenBank® Accession No. X65308 ). The pGEM®-T Vector was created by linearizing the pGEM®-5Zf(+) Vector with EcoRV at base 51 and adding a T to both 3´-ends. The position of the T is indicated by * in the pGEM®-T Vector Sequence (.txt).
In the pGEM®-T Vector, T7 and SP6 RNA polymerase promoters flank a multiple cloning region within the α-peptide coding region for β-galactosidase. Insertional inactivation of the α-peptide allows recombinant clones to be directly identified by Blue/White Screening on indicator plates. The vector allows preparation of single-stranded DNA due to its f1 Origin of Replication.
The multiple cloning site is flanked by recognition sites for the restriction enzyme BstZI, allowing release of the insert by a single-enzyme digestion. Alternatively, a double digestion may be used to release the insert from the vector.
The pGEM®-T Vector System II contains JM109 Competent Cells in addition to all of the pGEM®-T Vector System I components.
References
Mezei, L.M. and Storts, D.R. (1994) In: PCR Technology: Current Innovations , Griffin, H.G. and Griffin, A.M., eds., CRC Press, Boca Raton, FL, 21.
Robles, J. and Doers, M. (1994) Promega Notes 45 , 19–20.
Clark, J.M. (1988) Nucl. Acids Res . 16 , 9677–86.
Newton, C.R. and Graham, A. (1994) In: PCR , BIOS Scientific Publishers, Ltd., Oxford, UK, 13.