对从高盐稀态酱醪中筛选得到的米曲霉(Aspergillus oryzae)K9进行常压室温等离子体(atmospheric and room temperature plasma,ARTP)复合紫外诱变,并利用高盐含量平板进行驯化培养和筛选,旨在得到产耐盐高活性蛋白酶的菌株。首先对K9进...对从高盐稀态酱醪中筛选得到的米曲霉(Aspergillus oryzae)K9进行常压室温等离子体(atmospheric and room temperature plasma,ARTP)复合紫外诱变,并利用高盐含量平板进行驯化培养和筛选,旨在得到产耐盐高活性蛋白酶的菌株。首先对K9进行ARTP诱变,筛选得到优势菌株S-G27,其中性蛋白酶活性较K9提高35.82%;接着以S-G27为出发菌株进行紫外诱变,筛选得到优势菌株G-U2,其中性蛋白酶活性较K9提高56.81%;G-U2所产蛋白酶的耐盐性进一步提高,在15%NaCl溶液中,G-U2中性蛋白酶活性残留率为K9的1.94倍;G-U2稳定性良好,第7代时菌株的中性蛋白酶活性残留率为92%;和出发菌株K9相比,G-U2的菌丝更短、孢子更多、孢子体积更大且表面纹饰更多;诱变未引起黄曲霉毒素相关基因的表达,符合生产安全要求。在模拟发酵酱油实验中,发酵终点G-U2中性蛋白酶活性、酸性蛋白酶活性、氨基酸态氮质量浓度分别较K9高出120.19%、52.29%、18.37%。展开更多
Higher levels of acetaldehyde in beer are one of the major concerns in the current beer industry.Yeast produces acetaldehyde during alcoholic fermentation,and its modification significantly affects beer flavor and qua...Higher levels of acetaldehyde in beer are one of the major concerns in the current beer industry.Yeast produces acetaldehyde during alcoholic fermentation,and its modification significantly affects beer flavor and quality.A different mutant strain with lower acetaldehyde production and improved ethanol tolerance was constructed using the ARTP-ALE mutagenesis strategy with 4-methylpyrazole-disulfiram.As a result of the mutation,the alcohol dehydrogenase activity of the mutant strain decreased to about 71.22%of that of the wild-type strain.At the same time,the fermentation properties and genetic stability of the newly screened strain showed slight differences from the wild-type strain,and there were no safety concerns regarding industrial use of the mutant strain.展开更多
利用常温常压等离子体诱变(Atmospheric and Room Temperature Plasma, ARTP)蜡状芽孢杆菌,筛选获得酶活提升的菌株。结果表明,蜡状芽孢杆菌在种子液中培养6 h可达到对数期中后期,常温常压等离子体诱变时间为60 s时,蜡状芽孢杆菌的致死...利用常温常压等离子体诱变(Atmospheric and Room Temperature Plasma, ARTP)蜡状芽孢杆菌,筛选获得酶活提升的菌株。结果表明,蜡状芽孢杆菌在种子液中培养6 h可达到对数期中后期,常温常压等离子体诱变时间为60 s时,蜡状芽孢杆菌的致死率达到85%,筛选得到一株壳聚糖酶活性提高13.19%的突变株,然后进行酶活稳定性验证,经六代培养的平均酶活为9.643367 U/mL,波动在5%之内,表明其产酶量高,稳定性强,可用于后续出发菌株。诱变后菌株经电镜观察,发现菌落形态无明显变化,但是菌株的个体形态发生了变化,比原始菌株细长,证明酶活提高是ARTP诱变起到的作用。展开更多
L-丝氨酸在医药、食品和化工等领域应用广泛,然而L-丝氨酸是目前工业化生产难度较大的氨基酸之一。该文以实验室前期构建的产L-丝氨酸的谷氨酸棒杆菌A36-AB为出发菌株,利用常温常压等离子体诱变(atmospheric and room temperature plasm...L-丝氨酸在医药、食品和化工等领域应用广泛,然而L-丝氨酸是目前工业化生产难度较大的氨基酸之一。该文以实验室前期构建的产L-丝氨酸的谷氨酸棒杆菌A36-AB为出发菌株,利用常温常压等离子体诱变(atmospheric and room temperature plasma,ARTP)结合高通量筛选,获得突变菌株A36-AB-pDser-6。突变菌株前期生长迅速,60 h时L-丝氨酸产量为15.60 g/L。进一步利用响应面法优化突变菌株发酵培养基,L-丝氨酸产量达到29.23 g/L,生产强度为0.48 g/(L·h),生产强度较出发菌株[0.36 g/(L·h)]提高了33%;发酵周期从120 h缩短至60 h。对出发菌株及高产突变株进行全基因组测序及比较分析,发现突变株存在12个单碱基突变(single nucleotide variant,SNV)和15个插入缺失突变(insertions and deletion,InDel)。其中糖酵解途径中的关键酶葡萄糖-6-磷酸异构酶(由pgi基因编码)发生较多突变,这可能是由于突变菌株发酵周期缩短。该研究为利用谷氨酸棒杆菌产L-丝氨酸的工业化生产奠定了基础。展开更多
以实验室保藏的节杆菌属菌株Arthrobacter sp.SH为出发菌株,经常压室温等离子体(atmospheric and room temperature plasma,ARTP)诱变选育后,筛选到一株以淀粉为原料Tre Y-Tre Z途径生产海藻糖的高产突变菌株Arthrobacter sp.SH-52。该...以实验室保藏的节杆菌属菌株Arthrobacter sp.SH为出发菌株,经常压室温等离子体(atmospheric and room temperature plasma,ARTP)诱变选育后,筛选到一株以淀粉为原料Tre Y-Tre Z途径生产海藻糖的高产突变菌株Arthrobacter sp.SH-52。该突变菌株酶催化能力达到129.6 U/m L,比出发菌株提高了46.1%。对菌株Arthrobacter sp.SH-52的酶促反应条件进行优化,结果表明,酶促反应最适温度45℃,最适反应时间30 h,最适初始p H 6.5,最适底物为浓度20%的淀粉液化液(DE值为11.9)。经酶促反应优化后,采用未经纯化的粗酶液进行催化,转化率达到37.6%。实验结果表明ARTP诱变是选育高酶活海藻糖生产菌的有效方法,研究结果对工业化生产海藻糖具有一定的借鉴价值。展开更多
文摘对从高盐稀态酱醪中筛选得到的米曲霉(Aspergillus oryzae)K9进行常压室温等离子体(atmospheric and room temperature plasma,ARTP)复合紫外诱变,并利用高盐含量平板进行驯化培养和筛选,旨在得到产耐盐高活性蛋白酶的菌株。首先对K9进行ARTP诱变,筛选得到优势菌株S-G27,其中性蛋白酶活性较K9提高35.82%;接着以S-G27为出发菌株进行紫外诱变,筛选得到优势菌株G-U2,其中性蛋白酶活性较K9提高56.81%;G-U2所产蛋白酶的耐盐性进一步提高,在15%NaCl溶液中,G-U2中性蛋白酶活性残留率为K9的1.94倍;G-U2稳定性良好,第7代时菌株的中性蛋白酶活性残留率为92%;和出发菌株K9相比,G-U2的菌丝更短、孢子更多、孢子体积更大且表面纹饰更多;诱变未引起黄曲霉毒素相关基因的表达,符合生产安全要求。在模拟发酵酱油实验中,发酵终点G-U2中性蛋白酶活性、酸性蛋白酶活性、氨基酸态氮质量浓度分别较K9高出120.19%、52.29%、18.37%。
基金Supported by Heilongjiang Natural Science Foundation Joint Guide Project(LH2019C022)。
文摘Higher levels of acetaldehyde in beer are one of the major concerns in the current beer industry.Yeast produces acetaldehyde during alcoholic fermentation,and its modification significantly affects beer flavor and quality.A different mutant strain with lower acetaldehyde production and improved ethanol tolerance was constructed using the ARTP-ALE mutagenesis strategy with 4-methylpyrazole-disulfiram.As a result of the mutation,the alcohol dehydrogenase activity of the mutant strain decreased to about 71.22%of that of the wild-type strain.At the same time,the fermentation properties and genetic stability of the newly screened strain showed slight differences from the wild-type strain,and there were no safety concerns regarding industrial use of the mutant strain.
文摘利用常温常压等离子体诱变(Atmospheric and Room Temperature Plasma, ARTP)蜡状芽孢杆菌,筛选获得酶活提升的菌株。结果表明,蜡状芽孢杆菌在种子液中培养6 h可达到对数期中后期,常温常压等离子体诱变时间为60 s时,蜡状芽孢杆菌的致死率达到85%,筛选得到一株壳聚糖酶活性提高13.19%的突变株,然后进行酶活稳定性验证,经六代培养的平均酶活为9.643367 U/mL,波动在5%之内,表明其产酶量高,稳定性强,可用于后续出发菌株。诱变后菌株经电镜观察,发现菌落形态无明显变化,但是菌株的个体形态发生了变化,比原始菌株细长,证明酶活提高是ARTP诱变起到的作用。
文摘以实验室保藏的节杆菌属菌株Arthrobacter sp.SH为出发菌株,经常压室温等离子体(atmospheric and room temperature plasma,ARTP)诱变选育后,筛选到一株以淀粉为原料Tre Y-Tre Z途径生产海藻糖的高产突变菌株Arthrobacter sp.SH-52。该突变菌株酶催化能力达到129.6 U/m L,比出发菌株提高了46.1%。对菌株Arthrobacter sp.SH-52的酶促反应条件进行优化,结果表明,酶促反应最适温度45℃,最适反应时间30 h,最适初始p H 6.5,最适底物为浓度20%的淀粉液化液(DE值为11.9)。经酶促反应优化后,采用未经纯化的粗酶液进行催化,转化率达到37.6%。实验结果表明ARTP诱变是选育高酶活海藻糖生产菌的有效方法,研究结果对工业化生产海藻糖具有一定的借鉴价值。
