Expression of Genetically Engineered BL21(DE3) with the Staphylokinase Gene from Staphylococcus aureus
DOI:
https://doi.org/10.36320/ajb/v9.i3.7915Keywords:
Cloning, fibrinolytic property, isopropyl -D-1-thiogalactopyranoside, protein expression, staphylokinaseAbstract
The current study aimed to obtain staphylokinase (Sak) protein as a fibrinolytic agent from Escherichia coli as a nonpathogenic bacterium after transforming it with a cloned vector. From 630 clinical samples, 98 isolates of S. aureus were obtained and 81 (82.65%) were Sak producers. PCR amplification of the sak gene identified a single DNA band with the expected length of 408 bp in three S. aureus isolates. E. coli DH5a was successfully transformed with the sak gene. PCR amplification showed that the recombinant gene comprised 408 bp, which was the same size as the native gene amplified from S. aureus. Bacterial proteins of genetically engineered BL21(DE3) with cloned pET-28a-sak were separated according to their molecular weight and recombinant Sak appeared as a band with an estimated molecular weight ca 15.5 KDa. A non-induced sample with cloned vector was included as control. The fibrinolytic activity of recombinant Sak was higher than that of the native Sak. We found that the gene expression level and Sak production rate in the genetically engineered BL21(DE3) were similar to those of the native S. aureus.
Key words: Cloning, fibrinolytic property, isopropyl b-D-1-thiogalactopyranoside, protein expression, staphylokinase
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Copyright (c) 2017 Hêro Farhad Salah, Adel Kamal Khider, Sekaran Muniandy
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