pGEM-T Vector System II
Efficient PCR 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.
T7 and SP6 RNA polymerase promoters flank a multiple cloning region within the ?-peptide coding region for ?-galactosidase. Insertional inactivation of the ?-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.
- 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.
pGEM®-T and pGEM®-T Easy Vector Systems Technical Manual
PDF (434 KB)
pGEM T and pGEM T Easy Vector Systems FB033
PDF (202 KB)
Certificate of AnalysisLookup Certificate of Analysis
-30C TO -10C
For Research Use Only. Not for Use in Diagnostic Procedures.