The pGEM®-T and pGEM®-T Easy vector systems are used for cloning PCR products. The insert site at the 3’ end with a thymidine overhang improves the efficiency of PCR product ligation. These vectors contain T7 and SP6 RNA polymerase promoters and multiple cloning sites for recombinant cloning and generation of a series of nested deletions using the Erase-a-Base® system.
This manual provides detailed procedures for using the pGEM®-T and pGEM®-T Easy vector systems, including vector description, maps, component lists, ligation steps, transformation protocols, and troubleshooting. It also contains appendices with vector sequences, restriction sites, and reference formulations.
The document is organized into sections covering description, vector maps, product components, ligation with 2× rapid ligation buffer, transformation using high‑efficiency JM109 competent cells, precautions, plasmid DNA extraction, single‑strand DNA preparation, and a troubleshooting guide. Additional reference material and product catalogs are listed in the appendices. This manual is written in Chinese.
| Vector amount per reaction | 1.2 µg (50 ng/µl) |
|---|---|
| Reactions per kit | 20 |
| Storage temperature for competent cells | -70 °C |
Use JM109 high‑efficiency competent cells (catalog number L2001).
Dissolve 1.2 g IPTG in water and bring to 50 mL (0.1 M).