未脱毒水解液中的醛类、酚类等物质抑制菌株的生长发酵。产甘油假丝酵母(Candida glycerinogenes)具有多重抗逆性能,过表达相关抗性基因构建重组菌株并进行筛选。相比于对照菌株,过表达CgGCN4的重组菌株在含有3.0 g/L糠醛、2.0 g/L香草醛、3.5 g/L乙酸的高糖培养基中生物量分别提高25.2%、14.6%、5.2%;葡萄糖消耗速率分别提高9.9%、5.5%、13.1%;在3.0 g/L糠醛的胁迫下甘油产量提高23.4%。在糠醛、香草醛或乙酸的胁迫下,重组菌株胞内活性氧水平的提高程度分别降低35.2%、28.0%、15.3%,与碘化丙啶的结合水平均有不同程度的降低,表明重组菌株可通过降低胞内活性氧的积累、保持细胞膜的完整性来保持菌株优良的生长与发酵性能。将抗性重组菌株应用于未脱毒甘蔗渣水解液开放式发酵生产甘油,可使耗糖速率提高20.0%;生物量提高12.3%;甘油产量提高21.7%。该研究为未脱毒纤维素水解液的应用提供了新思路及遗传资源。
Aldehydes and phenols in the undetoxified hydrolysate inhibited growth and fermentation of the strain.Candida glycerinogenes have multiple resistance properties, and recombinant strains were constructed by overexpressing relevant resistance genes and screened.Compared with the control strain, the recombinant strain overexpressing CgGCN4 increased the biomass of C.glycerinogenes by 25.2%, 14.6%, and 5.2%, and the glucose consumption rate by 9.9%, 5.5%, and 13.1%, respectively, in a high-glucose medium containing 3.0 g/L furfural, 2.0 g/L vanillin and 3.5 g/L acetic acid, and increased glycerol production by 23.4% under the stress of 3.0 g/L furfural.Under the stress of furfural, vanillin or acetic acid, the increase of intracellular reactive oxygen species level of the recombinant strains was reduced by 35.2%, 28.0%, and 15.3%, respectively, and the binding level of propidium iodide was reduced to different degrees, suggesting that the recombinant strains could maintain the excellent growth and fermentation performance of the strains by reducing the accumulation of intracellular reactive oxygen species and maintaining the integrity of the cell membrane.Application of the resistant recombinant strain to open fermentation of undetoxified bagasse hydrolysate for glycerol production resulted in a 20.0% increase in sugar consumption rate, a 12.3% increase in biomass and a 21.7% increase in glycerol yield.This study provides new ideas and genetic resources for the application of undetoxified cellulose hydrolysate.
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