gTME构建共发酵木糖和葡萄糖的重组酿酒酵母

gTME for Construction of Recombinant Yeast Co-fermenting Xylose and Glucose

利用全转录工程(gTME)方法将全局转录因子spt15随机突变并克隆表达,构建突变库。将突变基因连接到表达载体pYX212上,醋酸锂法转化入不利用木糖的酿酒酵母YPH499中,经特定的培养基初筛获得高效利用木糖并共发酵木糖和葡萄糖的酿酒酵母重组菌株。对获得的重组菌株进行了初步研究,该菌株能够很好的利用木糖并共发酵木糖和葡萄糖。在30oC,200r/min,发酵96h时,50g/L木糖和葡萄糖的利用率为94.0%和98.9%,乙醇产率为32.4%和31.6%,原始菌株乙醇产率为44.3%;当木糖和葡萄糖以质量比1:1混合发酵时,木糖和葡萄糖利用率分别为91.7%和85.9%,乙醇产率为26%。木糖醇的含量极低。

Global transcription machinery engineering （gTME） was employed to engineer xylose metabolism. Mutation of the transcription factor gene Spt15 was introduced by error-prone PCR, followed by screening on media using xylose as the sole carbon source. One recombinant strain growing well on such media was chosen for further research. This strain showed modest growth rates in the media containing 50 g/L xylose or glucose at the condition of 30℃, 200 r/min, 96 h, 94.0% and 98.9% of xylose and glucose were consumed, with the ethanol yield were 32.4% and 31.6%, respectively. The control strain had the ethanol yield of 44.3% under the glucose concentration of 50 g/L. When the carbon source was 50 g/L glucose/xylose （1：1）, the utilization ratio of xylose and glucose was 91.7% and 85.9%, with the ethanol yield was 26%. Xylose was eventually exhausted. Concentration of the by-product xylitol was very low.