The Zero Blunt PCR Cloning Kit is designed to clone blunt PCR fragments (or any blunt DNA fragment) with a low background of non-recombinants. The pCR-Blunt vector contains the lethal E. coli ccdB gene fused to the C-terminus of LacZα.
Ligation of a blunt PCR fragment disrupts expression of the lacZα-ccdB gene fusion permitting growth of only positive recombinants upon transformation. Cells that contain non-recombinant vector are killed when the transformation mixture is plated.
Speed with Flexibility
With the addition of the ExpressLink T4 DNA Ligase into Zero Blunt Cloning Kits, ligation can now be performed at room temperature in only 5 minutes with reactions typically yielding >80% recombinants containing inserts.
The pCR-Blunt Vector Features:
- ccdB gene for positive selection
- Choice of kanamycin or Zeocin resistance for flexible antibiotic selection
- EcoR I sites flanking the PCR product insertion site for excision of inserts
- T7 promoter/primer site for in vitro RNA transcription and sequencing
- M13 forward and reverse primer sites for sequencing or PCR screening
Performance and Value
Zero Blunt Cloning Kits are available without competent cells (K2750-20 and K2750-40) or with One Shot TOP10 Chemically Competent E. coli (K2700-20 and K2700-40), in both 20 and 40-reaction sizes.
| Zero Blunt Cloning Kits | |
|---|---|
| Cloning efficiency | 95% (with control) |
| Time for ligation | 5 minutes |
| Temperature for ligation | Room Temperature |
| PCR Clean-up required? | No |
| Selection Antibiotic | Kanamycin & Zeocin |
| Kit Contents | |
| Vector | |
| Ligation Buffer | |
| Forward and reverse sequencing primers | |
| dNTPs | |
| Sterile Water | |
| Control DNA template | |
Resources
For Research Use Only. Not intended for any animal or human therapeutic or diagnostic use.