Cloning and sequence analysis of Bacillus thuringiensis gene fragments isolated by restriction digest PCR

To clone and analyze Bacillus thuringiensis gene fragments isolated by restriction digest PCR (RD-PCR).

Specific primers were designed to amplify the genes of Bacillus thuringiensis israelensis (Bti), and the PCR products were classified and re-amplified by RD-PCR to obtain the fragments for subsequent purification and cloning into the pMD18-T vectors, followed by rapid identification. The recombinant plasmids were extracted from positive clones and the target gene fragments were sequenced.

Sequence analysis showed that all the fragments amplified were Bti genes.

RD-PCR is reliable in breaking down large gene fragments into confined and shorter gene fragments for preparing microarray probes.