摘要：
In order to construct Saccharomyces cerevisiae with α-amylase activity and low production of ethyl carbamate(EC).The expression vectors containing promotor sequence of ADH1 and ADH2 and α-amylase encoding sequence of Saccharomycopsis fibuligera were constructed,respectively.The α-amylase expression vectors were transformed into S△car1 and the positive recombinants were screened.The results indicated that the positive recombinants with α-amylase activity and reduced ethyl carbamate(EC) production were constructed and named S△car1/A1and S△car1/A2,respectively.In solid and liquid medium,α-amylase expression of S△car1/A1 was induced by glucose.Meanwhile,with the increasing of glucose concentration,the α-amylase expression was increased.However,the α-amylase of S△car1/A2 could be expressed without glucose and it could be feedback-inhibited by glucose.The results indicated α-amylase expression of S△car1/A2 was increased by low glucose concentration and reduced by high glucose concentration,although its expression was higher in the presence of glucose than without glucose.In conclusion,the ability of S△car1/A2 to produce α-amylase and hydrolyze starch was higher than S△car1/A1.In view of those,the S△car1/A2 could be used to ferment the materials containing starch.However,further research should be carried out about the condition of enzyme expression,catalyzing and fermentation.%为了构建可利用淀粉而低产氨基甲酸乙酯的酵母菌株,通过构建分别含ADH1和ADH2启动子的α-淀粉酶表达载体pADH1/alp1和pADH2/alp1,并将其分别转入精氨酸酶基因缺失的酵母菌株S△car1中.在此基础上,分析比较其在固态和液态条件下产α-淀粉酶的能力以及葡萄糖对α-淀粉酶表达的影响.结果表明:可利用淀粉而低产氨基甲酸乙酯的黄酒酵母S△car1/A1和S△car1/A2已成功构建.S△car1/A1在固体和液体培养基中产生淀粉酶时,都需要葡萄糖的诱导,且随着培养基中葡萄糖浓度的增加,酶的表达量增加.S△car1/A2在两种培养基中产生淀粉酶时,不需葡萄糖诱导,但S△car1/A2 α-淀粉酶表达受葡萄糖的反馈抑制,低浓度的葡萄糖会促进α-淀粉酶的产生,过高的葡萄糖浓度使其α-淀粉酶的表达量减少,但是在含葡萄糖的培养基中的产酶量要比在不含葡萄糖的培养基上的高.总体而言,S△car1/A2产α-淀粉酶和水解淀粉的能力比S△car1/A1的强.因此,S△car1/A2可被用于生料发酵,但对其在实际生产中的产酶、催化和发酵条件等则有待进一步研究.
