title: Engineering a wild-type diploidSaccharomyces cerevisiaestrain for second-generation bioethanol production.
abstract: BACKGROUND
  The cost-effective production of second-generation bioethanol, which is made from lignocellulosic materials, has to face the following two problems: co-fermenting xylose with glucose and enhancing the strain's tolerance to lignocellulosic inhibitors. Based on our previous study, the wild-type diploidSaccharomyces cerevisiaestrain BSIF with robustness and good xylose metabolism genetic background was used as a chassis for constructing efficient xylose-fermenting industrial strains. The performance of the resulting strains in the fermentation of media with sugars and hydrolysates was investigated.
  
  
  RESULTS
  The following two novel heterologous genes were integrated into the genome of the chassis cell: the mutantMGT05196N360F, which encodes a xylose-specific, glucose-insensitive transporter and is derived from theMeyerozyma guilliermondiitransporter geneMGT05196, and Ru-xylA(where Ru represents the rumen), which encodes a xylose isomerase (XI) with higher activity inS. cerevisiae. Additionally, endogenous modifications were also performed, including the overproduction of the xylulokinase Xks1p and the non-oxidative PPP (pentose phosphate pathway), and the inactivation of the aldose reductase Gre3p and the alkaline phosphatase Pho13p. These rationally designed genetic modifications, combined with alternating adaptive evolutions in xylose and SECS liquor (the leach liquor of steam-exploding corn stover), resulted in a final strain, LF1, with excellent xylose fermentation and enhanced inhibitor resistance. The specific xylose consumption rate of LF1 reached as high as 1.089 g g-1h-1with xylose as the sole carbon source. Moreover, its highly synchronized utilization of xylose and glucose was particularly significant; 77.6% of xylose was consumed along with glucose within 12 h, and the ethanol yield was 0.475 g g-1, which is more than 93% of the theoretical yield. Additionally, LF1 performed well in fermentations with two different lignocellulosic hydrolysates.
  
  
  CONCLUSIONS
  The strain LF1 co-ferments glucose and xylose efficiently and synchronously. This result highlights the great potential of LF1 for the practical production of second-generation bioethanol.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Shan Da Nan Road 27, Jinan, 250100 China.|Shandong Provincial Key Laboratory of Microbial Engineering, Qi Lu University of Technology, Jinan, 250353 China.
authors.forename: Hongxing
authors.initials: H
authors.lastname: Li
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Shan Da Nan Road 27, Jinan, 250100 China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Shen
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Shan Da Nan Road 27, Jinan, 250100 China.
authors.forename: Meiling
authors.initials: M
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Shan Da Nan Road 27, Jinan, 250100 China.
authors.forename: Jin
authors.initials: J
authors.lastname: Hou
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Shan Da Nan Road 27, Jinan, 250100 China.
authors.forename: Chunlei
authors.initials: C
authors.lastname: Jiao
authors.affiliation: Shandong Provincial Key Laboratory of Microbial Engineering, Qi Lu University of Technology, Jinan, 250353 China.
authors.forename: Zailu
authors.initials: Z
authors.lastname: Li
authors.affiliation: Shandong Provincial Key Laboratory of Microbial Engineering, Qi Lu University of Technology, Jinan, 250353 China.
authors.forename: Xinli
authors.initials: X
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Shan Da Nan Road 27, Jinan, 250100 China.|Shandong Provincial Key Laboratory of Microbial Engineering, Qi Lu University of Technology, Jinan, 250353 China.
authors.forename: Xiaoming
authors.initials: X
authors.lastname: Bao
keywords: Budding yeast; Lignocellulosic ethanol; Synchronous utilization of xylose and glucose; Xylose isomerase; Xylose-specific transporter