Cloning and expression of metallothionein mutant α-KKS-α in Anabaena sp. PCC 7120

The mouse metallothionein (mMT) mutant α-KKS-α has a higher capacity for binding heavy metals than wild type mMT. The mMT mutant α-KKS-α gene was placed under the control of the strong promoter PpbsA to generate the intermediate vector pRL-α-KKS-α. pRLα-KKS-α was then linked with the plasmid pDC-08 to construct shuttle expression vector pDC-αKKS-α. This expression vector was transformed into Anabaena sp. PCC 7120 using triparental conjugative transfer. After antibiotic selection (ampicillin and kanamycin), transgenic Anabaena was identified by PCR and Western blotting. The expression level of the mMT mutation α-KKS-α reached 7.4 mg/g dry cells weight, as detected by ELISA, and heavy metal resistance of the transgenic Anabaena was significantly improved.