CloneJET PCR 克隆试剂盒

The pJET1.2 vector is not specifically designed for protein expression. It contains the T7 RNA promoter for in vitro transcription of the cloned insert. In some cases, this T7 promoter can be used for non-toxic gene expression in bacterial strains having IPTG inducible T7 RNA polymerase (like E. coli BL21); however, the promoter does not contain regulatory elements and protein expression will not be well regulated.

Using pJET1.2 Forward and Reverse Sequencing Primers in PCR (as described in the protocol), negative colonies would yield PCR products that are about 120 bp long. Positive colonies would yield longer PCR products that should reflect the insert size (with addition of 120 bp from the vector).

The insert can be expected at the Eco32I (EcoRV) site in the vector at the position 369 since the pJET1.2/blunt cloning vector was linearized with Eco32I.

Any DNA fragment (either blunt- or sticky-end) that is 6 bp to 10 kb long and generated from PCR or restriction digestion, can be successfully cloned using the kit.

Phosphorylation of the PCR primers is not required since the 5'-ends of the pJET1.2/blunt vector contain phosphoryl groups for ligation.