Construction of secretory expression vector of Trichoderma reesei and expression of enhanced GFP

Abstract:

[Objective] To construct a secretory expression vector of Trichoderma reesei and prove its feasibility by recombinant expression of enhanced green fluorescence protein (eGFP), and observe the process of the expression of eGFP, spontaneously. [Methods] Construct the expression vector pPth15 by successively ligate the promoter of the CBH1 gene of T. reesei, the signal peptide sequence, the terminator and the hygromycin resistance gene into the backbone plasmid pUC19. Insert eGFP gene into pPth15 to aquire the recombinant plasmid pPth15-eGFP, and transformate it into the T. reesei protoplasts. Screening positive transformants via hygromycin resistance and PCR amplification. [Results] After culturing the positive transformants on the PDA medium for 2 to 3 days, green fluorescence distribution was emerged on the hyphae tips, the septum and the extracellular medium. [Conclusion] The constructed expression vector could be used for eGFP expression, and this work makes contribution to the follow-up study of heterogeneous protein expression in T. reesei and provides a reference in the research of extracellular protein expression of T. reesei.