All components of the Lethal Based Fast Cloning Kit should be stored at -20°C. Repeated freeze-thaw should be avoided.

The Lethal Based Fast Cloning Kit is an advanced positive clone selection system for the highest efficiency cloning of PCR products and any other DNA fragment, either blunt or sticky-end. This kit works for both phosphorylated and non-phosphorylated DNA fragments. By using this kit, the whole process of positive clone selection and ligation takes only 5 minutes and yields more than 95% positive clones. Blunt end PCR products amplified by DNA polymerases which have proofreading activity (e.g., Pfu polymerase) can be inserted directly into vector. PCR products that amplified by DNA polymerases which don’t have proofreading activity (e.g., Taq polymerase) need to be treated with Blunting Enzyme in this kit (7 min) before ligation. Ligation products can be used on the transformation of common used E. coli strains.

The kit features a novel positive selection cloning vector pLB vector. This vector contains a lethal gene (contains multiple cloning sites) which would be disrupted by the insertion of a DNA fragment into the cloning site. As a result, only cells with recombinant plasmids are able to propagate, cyclized empty pLB Vector will express lethal toxic protein which will kill the E. coli cells. This cloning method eliminates the need of expensive blue/white screening and accelerates the process of cloning and selection.

■ Quick and efficient: Blue/white screening is not necessary, and the whole ligation process can be finished within 5 min.

■ Sensitive and widely applied: Good for the ligation of low concentration fragments and long fragments, works for both blunt and sticky end product.