Cloning, expression and biological activity identification of Cu-Zn SOD from Trichoderma harzianum

To explore the integrated bio-control mechanism of Trichoderma harzianum and acquire some bio-control associated novel genes, a Trichoderma harzianum mycelium cDNA library has been constructed and thereby randomly selected clones were sequenced and analyzed by bioinformatics analysis. Full-length cDNA encoding Cu-Zn SOD was successfully cloned. The results indicated that the ORF of Cu-Zn SOD was 465 bp, 154 amino acids were encoded, and the deduced molecular weight was 15.7 ku. The gene was ligated to the vector of pET28 and thereby transformed to the E.coli BL21. After screening the correct recombinant E.coli pET28a-SOD/BL21 transformants, it was expressed by IPTG and the expressed products were analyzed with SDS-PAGE. The expression condition was optimized to obtain the maximum quantity of SOD. And then the protein activity was detected by SOD kit. The optimization condition for SOD expression was as follows: cultivating at 37°C and initial induction at OD₆₀₀ of 0.6 with IPTG of 0.125 mmol/L for 4 h. The protein activity was 26.69 U/mL. The SOD gene was expressed successfully and the recombinant protein was obtained. This study provides the foundation for studying the molecular mechanism of Trichoderma harzianum in future research.