Effect of BAT2 Deletion with Lg-ATF1 Overexpression on the Production of Higher Alcohols and Ester in Brewer’s Yeast
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Abstract:
Ethyl acetate and higher alcohols are important flavor substances in beer. The effect of BAT2 gene and Lg-ATF1 gene in beer yeast on producing alcohol esters was explored to solve the problem of more higher alcohols and less ethyl acetate in beer. The expression vector pUC-PLABBK was constructed by using the method of enzyme digestion and connection. Saccharomyces cerevisiae multiploid S5 was used as the original strain and the KanMX gene was used as a selection marker in this study. Lithium acetate transformation and homologous recombination were employed to screen for a mutant strain S5-Lg that overexpressing Lg-ATF1 and at the same time deleting BAT2. The variation between the content of higher alcohols, ethyl acetate and expression of related genes in the engineered yeast strain S5-Lg was investigated by fermentation experiments and digital PCR. The results showed that compared with the original strain S5, the ethyl acetate production of the S5-Lg strain was increased by 26.81%, total higher alcohol production decreased by 45.72%, of which the contents of isobutanol and isoamyl alcohol were reduced by 10.63% and 9.55%, respectively. The expression of Lg-ATF1 gene was greatly increased, and the expression level of BAT2 gene was decreased by 45.72%. It effectively changed the ability of brewer's yeast to produce ethyl acetate and higher alcohols and has important significance for improving beer flavor.
