We report the expression profile of acyl-lipid △12-desaturase （desA） gene from Synechocystis sp. PCC6803 and its effect on cell membrane lipid composition and cold tolerance in prokaryotic （Escherichia coil） and ...We report the expression profile of acyl-lipid △12-desaturase （desA） gene from Synechocystis sp. PCC6803 and its effect on cell membrane lipid composition and cold tolerance in prokaryotic （Escherichia coil） and eukaryotic （Solanum tuberosum） cells. For this purpose, a hybrid of desA and reporter gene encoding thermostable lichenase （licBM3） was constructed and used to transform these cells. The expression of this hybrid gene was measured using qualitative （Petri dish test, electrophoregram and zymogram） and quantitative methods （spectrometry and gas liquid chromatography assays）. The maximum level of linoleic acid in the bacterial cells containing hybrid gene was 1.9% of total fatty acids. Cold stress tolerance assays using plant damage index and growth parameters showed that cold tolerance was enhanced in primary transgenic lines because of increased unsaturated fatty acid concentration in their lipids. The greatest content of 18：2 and 18：3 fatty acids in primary transgenic plants was observed for lines 2 （73%） and 3 （41%）. Finally, our results showed that desaturase could enhance tolerance to cold stress in potato, and desaturase and lichenase retain their functionality in the structure of the hybrid protein where the enzymatic activity of target gene product was higher than in the case of reporter lichenase gene absence in the construction.展开更多
本试验根据GenBank上登录的牛瑟氏泰勒虫ITS基因序列(AY661522.1),应用Primer Premier 5.0和Oligo 6.31软件设计合成1对特异性引物,以牛瑟氏泰勒虫DNA为模板,建立了牛瑟氏泰勒虫ITS基因PCR诊断方法。该方法扩增片段大小为1020 bp,与参...本试验根据GenBank上登录的牛瑟氏泰勒虫ITS基因序列(AY661522.1),应用Primer Premier 5.0和Oligo 6.31软件设计合成1对特异性引物,以牛瑟氏泰勒虫DNA为模板,建立了牛瑟氏泰勒虫ITS基因PCR诊断方法。该方法扩增片段大小为1020 bp,与参考序列的同源性为98%;建立的PCR方法与猪附红细胞体、犬新孢子虫和弓形虫均无交叉反应,最低DNA检出量为1.5 pg/μL;通过对60份临床样品的检测,并与血涂片方法进行比较,结果显示PCR方法具有特异、敏感等特点,适用于牛瑟氏泰勒虫的检测。展开更多
目的:建立食品中阪崎肠杆菌双重PCR检测方法。方法:以食品中污染的阪崎肠杆菌为研究对象,以其基因组16 S rRNA-23 S rRNA的内部转录间隔区(ITS)基因和外膜蛋白A(OmpA)基因为靶标选择引物,进行双重PCR扩增,产物用琼脂糖凝胶电泳检测。结...目的:建立食品中阪崎肠杆菌双重PCR检测方法。方法:以食品中污染的阪崎肠杆菌为研究对象,以其基因组16 S rRNA-23 S rRNA的内部转录间隔区(ITS)基因和外膜蛋白A(OmpA)基因为靶标选择引物,进行双重PCR扩增,产物用琼脂糖凝胶电泳检测。结果:在优化的双重PCR条件下,阪崎肠杆菌的ITS基因和OmpA基因均可得到有效扩增,方法的灵敏度为1.6×102cfu/mL。所建立的方法用于实际食品样品检测,结果稳定。结论:本研究所建立的方法特异好、灵敏较高、分析周期短、结果准确,可用于食品中阪崎肠杆菌的日常检测。展开更多
基金supported by the program "Biological Diversity–Gene Foundation and Genetic Diversity" of the Presidium of the Russian Academy of Sciences and the Russian Foundation for Basic Research (project nos. 08-04-90410-Ykp-a, 05-04-49186-a, 04-04-81039-Bel_a)
文摘We report the expression profile of acyl-lipid △12-desaturase （desA） gene from Synechocystis sp. PCC6803 and its effect on cell membrane lipid composition and cold tolerance in prokaryotic （Escherichia coil） and eukaryotic （Solanum tuberosum） cells. For this purpose, a hybrid of desA and reporter gene encoding thermostable lichenase （licBM3） was constructed and used to transform these cells. The expression of this hybrid gene was measured using qualitative （Petri dish test, electrophoregram and zymogram） and quantitative methods （spectrometry and gas liquid chromatography assays）. The maximum level of linoleic acid in the bacterial cells containing hybrid gene was 1.9% of total fatty acids. Cold stress tolerance assays using plant damage index and growth parameters showed that cold tolerance was enhanced in primary transgenic lines because of increased unsaturated fatty acid concentration in their lipids. The greatest content of 18：2 and 18：3 fatty acids in primary transgenic plants was observed for lines 2 （73%） and 3 （41%）. Finally, our results showed that desaturase could enhance tolerance to cold stress in potato, and desaturase and lichenase retain their functionality in the structure of the hybrid protein where the enzymatic activity of target gene product was higher than in the case of reporter lichenase gene absence in the construction.
文摘目的:建立食品中阪崎肠杆菌双重PCR检测方法。方法:以食品中污染的阪崎肠杆菌为研究对象,以其基因组16 S rRNA-23 S rRNA的内部转录间隔区(ITS)基因和外膜蛋白A(OmpA)基因为靶标选择引物,进行双重PCR扩增,产物用琼脂糖凝胶电泳检测。结果:在优化的双重PCR条件下,阪崎肠杆菌的ITS基因和OmpA基因均可得到有效扩增,方法的灵敏度为1.6×102cfu/mL。所建立的方法用于实际食品样品检测,结果稳定。结论:本研究所建立的方法特异好、灵敏较高、分析周期短、结果准确,可用于食品中阪崎肠杆菌的日常检测。
基金This research was funded by the grant fromthe National Natural Science Foundation of China (30499341 ,30270221) the Excellent YouthFoundationin Anhui Province (04043050)+2 种基金the Natural Science Foundation of Educational Committee of Anhui Province (2004sys003) the Foundation of Provincial Key Laboratory of Conservation and Utilizationfor I mportant Biological Resourcein Anhui the Special Foun-dation of Anhui Normal University
