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| Clone of Cu/Zn-Superoxide Dismutase Gene from Kluveromces lactis and Its expreesion in Saccharomyces cerevisiae |
| LI Wen-feng1,2,JI Jing1,WANG Gang1,NIU Bao-long1,WANG Hai-yong1,2 |
1.School of Agriculture and Bioengineering, Tianjin University, Tianjin 300072, China
2.School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China |
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Abstract According to Genbank Kluyveromyces lactis Cu/Zn-SOD gene sequence primers were designed；the Cu / Zn-SOD gene were obtained by PCR amplification. Driven by the PGK1 promoter, the gene fused to the fluorescent reporter gene GFP were used to construct recombinant plasmid YEplac195-PSGA and YCplac33-PSGA, and transformed into yeast (Saccharomyces cerevisiae) W303α strain. By colony PCR and fluorescence microscopic observation we confirmed that the Cu / Zn-SOD gene of Kluyveromyces lactis was successfully expressed in W303α strain.20 mM paraquat were added to the positive transformants before fermentation. SOD activity and total activity were respectively 6.7 times and 4.7 times as high as those without paraquat in the biomass of the fermentation medium. To further investigate the impact of the Cu / Zn-SOD gene on the host sod1Δ strain EG118, heat shock treatment was applied. Results show that heat strikes capability of host tolerance in the following order of EG118 (YEplac195-PSGA)> EG118 (YCplac33-PSGA)> EG118. The results is not only necessary for the fermentation industry to prevent bacterial fermentation in the aging strains and furthermore, an enhanced capacity to provide some theoretical guidance, but also a foundamental of in vitro directed evolution of the Cu/Zn-SOD.
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Received: 12 April 2010
Published: 25 August 2010
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