Conclusion

Combined with our previous study, a total of four EGT synthetases were mined in P. eryngii , which were PeEgt1 , PeEgt2a ,PeEgt2b , and PeEgt2c . Using CRISPR/Cas9 technology, theEgt1 and Egt2 genes of P. eryngii were integrated into the genome of S. cerevisiae IMX581, and eight engineered strains of S. cerevisiae were successfully constructed, including four single-gene engineered strains: IMX581-PeEgt1 , IMX581-PeEgt2a , IMX581-PeEgt2b , IMX581-PeEgt2c ; three double-gene engineered strains: IMX581-PeEgt1 -2a , IMX581-PeEgt1 -2b , IMX581-PeEgt1 -2c ; one triple-gene engineered strain: Imx581-PeEgt1 -2a -2b . It has been found that EGT can be generated when only the PeEgt1 expression box is integrated; on this basis, the integration of PeEgt2a or PeEgt2bincreased EGT production by 53.61% and 63.50%, respectively, compared with single-gene engineered strain. In this study, the EGT synthase genes mined from P. eryngii was used to construct the synthesis pathway of EGT in S. cerevisiae and verify its function, which laid a foundation for further improving the synthesis pathway of EGT in edible fungi and providing effective genes for the construction of engineered strains with high expression of EGT.