Expression of chimeric chitinase 42 enzyme containing ChBD in its C-terminal in tobacco

One of the enzymes with vast potential in the biotechnology field is chitinase, which causes destruction of chitin. The current uses of chitinase include production of N-acetylglucosamine, isolation of protoplast from fungi and yeasts, preparation of monocellular proteins, control of plant pathogenic fungi, control of insect pests in the industries, especially the sugarcane industry, control of the transmission of malaria and as an additive used in antifungal creams and lotions. Chimeric chitinase 42 containing introns and also ChBD in its C-terminal was cloned into a eukaryotic expression vector pARM2 including His tag sequence. The His tag can be used for protein purification. The cloning was confirmed by PCR and digestion pattern using appropriate restriction enzymes. This expression construct used for transformation of tobacco using agrobacterium tumefaciens and A.rhizogenes. Tobacco plants and also hairy roots were regenerated on kanamycine medium. Gene integration was confirmed by PCR method using specific chit 42 primers. Transgenic events were checked by Vir G primer to verify that the amplified band is not agrobacterium born. Chit 42 expression was confirmed by Sodium Dodecyl Sulfate PolyAcrylamide Gel Electrophoresis (SDS-PAGE), protein dot blot and western blot.

کلیدواژه ها:

chimeric chitinase 42 enzyme ، His-tag sequence ، expression