Cloning and expression of lipase-subtilisin protein fusion

Autor: Mohammad Hasan Shahhosseini, Davoud Esmaeili, Neda Gharagozloo Hesari, Taher Mohammadian, Atousa Ferdosi
Rok vydání: 2020
Předmět:
Zdroj: Gene Reports. 20:100727
ISSN: 2452-0144
Popis: Wastewater production has also increased due to increasing population growth and increasing water consumption in communities. This increase in sewage production can be a major environmental hazard. Hospital wastewater is one of the types of wastewater that poses many hazards to the environment and is classified as hazardous by its quality. Therefore, essential to the output status hospital wastewater should be accurate and prevent hospital wastewater from entering without purifying the living environment. The aim of this study was the design and synthesis construct containing the lipase-subtilisin fusion gene in the plasmid pet22b vector. The recombinant plasmid then was transformed and expressed in expression host E. coli BL21DE3. Construct PET22b-LIP-SUB was designed by researchers and optimized with bioinformatics software. It was then synthesized by a Biometrics company. The transformation was performed and was induced in the presence of 1 mM concentration IPTG. For confirmation of recombinant protein SDS-PAGE and western blotting were performed. Nickel column was used for purification of the recombinant protein. Based on bioinformatics analysis, the recombinant protein had good spatial structure and stability and half-life. The recombinant protein was confirmed by SDS-PAGE and western blotting with Anti-his tag. The concentration of the recombinant protein was 4.11 mg/ml. The results of this research showed that recombination protein LIP-SUB contains heat and acid resistant and higher potency, therefore, require to evaluate biological activity in hospital wastewater.
Databáze: OpenAIRE
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