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Isolation of a fibrinolytic enzyme BKII gene from local isolate of Bacillus

Abstr Regulation and production of Fibrinolytic enzymes from bacterial sources especially from Bacillus strains has taken a leading role in the medical sciences for the treatment of cardiovascular disorders as it removes thrombus or clots adding to its significant role in curing human health issues saving millions. Significant progress has been made during the last few years on the studies of fibrinolytic enzymes in identifying, cloning, purification, characterization and overproduction of these for commercialization in medical sciences and in fields like detergents development. In our present research an act was made to isolate and clone a metalloprotease fibrinolytic enzyme Bacillokinase (BKII) gene from a local isolate of Bacillus spp. This enzyme has strong fibrinolytic activity and also reported to be plasminogen activator. Doubos salts media with a pH of 7.2 at 40 o C was employed for the maintenance of growth of locally isolated specie of Bacillus. PCR amplification of fibrinolytic enzyme gene was done using specifically designed primers from genomic DNA isolated from the bacterial cell culture under optimized conditions. The amplified product of 1381 base pairs including the peptide signal sequence along with ORF was then ligated into pTZ 57 R/T cloning vector and transformed in E. coli Top10 cells. The clone was then confirmed by restriction analysis. This study can further help in the study of genetic diversity of these enzymes, their characteristics, regulation and hyper production for its different applications.