Development of an inducible cellobiohydrolase promoter and its application for enhancing the production of ganoderic...

Ganoderma lingzhi, a well-known medicinal mushroom, is regarded as a promising cell factory for producing various high-value bioactive compounds. However, the lack of inducible genetic tools in Ganoderma has hindered metabolic and genetic engineering efforts in this species. In this study, the cellobiohydrolase gene promoter (Pcbh) from G. lingzhi was cloned for the first time. Using a codon-optimized yellow fluorescent protein gene as a reporter, G. lingzhi Pcbh showed the highest inducibility under Avicel induction, with greater mRNA accumulation and higher protein levels compared to induction by cellobiose or lactose. These findings were further confirmed using a codon-optimized Vitreoscilla hemoglobin gene as a reporter. Moreover, an induction regime containing 1.7% Avicel as the inducer and 1.8% glucose as an additional carbon source maintained high induction efficiency of G. lingzhi Pcbh without impairing cell growth. Furthermore, Pcbh was successfully applied to conditionally silence the lanosterol 14-α-demethylase gene (erg11), thereby enhancing the production of ganoderic acids (GAs) in G. lingzhi. In si-erg11 transformants, the maximum contents of GA-T, GA-S, GA-Me reached 43.78 ± 1.25, 71.7 ± 2.16, and 28.06 ± 1.30 μg/100 mg dry cell weight, respectively, representing 1.91-, 2.02-, and 1.91-fold increases over those of the control strain.