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To enhance the ethanol production efficiency of Saccharomyces cerevisiae in inulin fermentation, this study investigated the combined strategy of optimizing the secretion pathway and constructing diploid strains to improve the secretion efficiency of sucrose invertase（Suc2）and ethanol yield. Starting with the BY4741 and BY4742 strains, a PGK1p promoter replacement strategy was applied to construct recombinant strains BYCO and BY2CO, which overexpress the vesicle transport-related genes SEC1 and SSO2. Diploid strain BY3CO was obtained through diploid mating. Following the transformation of the engineered strains with an MFα signal peptide-tagged Suc2 expression vector, the transcription levels of SEC1 and SSO2, extracellular protein secretion, and Suc2 secretion efficiency were assessed. The results showed significant upregulation of SEC1 and SSO2 transcription in the recombinant strains. The extracellular protein secretion of the diploid strain BY3COmS increased by 216.67% compared to the original strain BYmS, and Suc2 secretion efficiency improved by 86.46%. Continuous fermentation analysis revealed that after 84 h, BY3COmS achieved a biomass of 35.00 g/L and ethanol production of 100.38 g/L, representing increases of 33.49% and 12.61%, respectively, compared to BYmS. Additionally, the inulin conversion rate improved by 10.32%, reaching 94.77%. This study demonstrates that the synergistic enhancement of vesicle transport pathways and the construction of diploid strains significantly improve Suc2 secretion efficiency and ethanol production performance in S. cerevisiae. These findings provide a novel technical approach for the development of efficient inulin-converting S. cerevisiae strains for bioethanol production.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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