[1]侯永杰),李杰),李庆华),等.杜氏盐藻寡糖基转移酶亚基stt3a酵母双杂交诱饵载体的构建及自激活和毒性检测*[J].郑州大学学报(医学版),2011,(06):835.
 HOU Yongjie),LI Jie),LI Qinghua,et al.Construction of yeast twohybrid bait plasmid for stt3a of oligosaccharyltransferase from Dunaliella salina and detection of selfactivation and toxicity of stt3a[J].JOURNAL OF ZHENGZHOU UNIVERSITY(MEDICAL SCIENCES),2011,(06):835.
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杜氏盐藻寡糖基转移酶亚基stt3a酵母双杂交诱饵载体的构建及自激活和毒性检测*
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《郑州大学学报(医学版)》[ISSN:1671-6825/CN:41-1340/R]

卷:
期数:
2011年06期
页码:
835
栏目:
专题研究
出版日期:
2011-11-20

文章信息/Info

Title:
Construction of yeast twohybrid bait plasmid for stt3a of oligosaccharyltransferase from Dunaliella salina and detection of selfactivation and toxicity of stt3a
作者:
侯永杰1)李杰1)李庆华1)王建人2)薛乐勋1)#
郑州大学生物工程系细胞生物学研究室 郑州 450005
Author(s):
HOU Yongjie1)LI Jie1)LI Qinghua1)WANG Jianren2)XUE Lexun1)
1)Laboratory for Cell Biology,Department of Biology, Zhengzhou University,Zhengzhou 450001 2)Department of Physiology, Henan College of Traditional Chinese Medicine,Zhengzhou 450008
关键词:
杜氏盐藻stt3a酵母双杂交鞭毛再生
Keywords:
Dunaliella salinastt3a geneyeast twohybridflagella regeneration
分类号:
Q781
摘要:
目的:观察杜氏盐藻寡糖基转移酶亚基stt3a的酵母双杂交诱饵载体表达产物对酵母细胞有无毒害作用并检测其自激活作用。方法:应用PCR方法获得stt3a可溶端的基因片段,将基因片段按正确的方向插入到酵母表达质粒pGBKT7中,经限制性内酶切鉴定正确后,用PEG/LiAc法转化到酵母菌株Y187中,并通过表型筛选检测诱饵蛋白有无毒性和自激活作用。结果:成功获得了stt3a可溶端的基因片段,其表达的蛋白对酵母菌株Y187无毒害,报告基因半乳糖苷酶活性没有被诱导。结论:酵母双杂交GAL4系统可以被用来研究杜氏盐藻中与stt3a相互作用的蛋白。
Abstract:
Aim: To observe whether expression products of oligosaccharyltransferase subunit stt3a bait plasmid affect the growth of yeast cells and its selfactivation.Methods:The cDNA fragments of stt3a were obtained by PCR and then inserted into the plasmid pGBKT7 to construct a recombinant bait plasmid pGBKT7stt3a.Subsequently, the bait plasmid was transformed into the yeast strain Y187 by PEG/LiAc method.Results:The cDNA fragments of stt3a were amplified successfully. All expression productions were not toxic to Y187 cells and activity of the reporter gene βgalactosidase was not induced.Conclusion:Yeast twohybrid GAL4 system can be utilized to study proteins interacted on stt3a in Dunaliella salina.

参考文献/References:

[1]Frank J,KaulfürstSoboll H,Rips S,et al.Comparative analyses of Arabidopsis complex glycan1 mutants and genetic interaction with staurosporin and temperature sensitive3a[J].Plant Physiol, 2008,148(3):1354 [2]Schaewen A,Frank F,Koiwa H.Role of complex Nglycans in plant stress tolerance[J].Plant Signal Behav,2008, 3(10): 871 [3]Chapman A,Li E,Kornfeld S.The biosynthesis of the major lipidlinked oligosaccharide of Chinese hamster ovary cells occurs by the ordered addition of mannose residues[J].J Biol Chem,1979,254(20):10243 [4]Koiwa H, Li F, McCully MG, et al. The STT3a subunit isoform of the Arabidopsis oligosaccharyltransferase controls adaptive responses to salt/osmotic stress[J]. Plant Cell, 2003,15(10):2273 [5]王翠,李杰,柳丽平,等.杜氏盐藻寡糖基转移酶亚基STT3a功能结构域的克隆与表达分析[J].生物工程学报,2010,26(6):760 [6]Igura M,Maita N,Kamishikiryo J,et al.Structureguided identification of a new catalytic motif of oligosaccharyltransferase[J]. EMBO J,2008,27(1):234 [7]Kim H, von Heijne G, Nilsson I. Membrane topology of the STT3 subunit of the oligosaccharyl transferase complex[J]. J Biol Chem,2005,280(21):20261 [8]Yan Q,Lennarz WJ.Studies on the function of oligosaccharyltransferase subunits,Stt3p is directly involved in the glycosylation process[J]. J Biol Chem,2002,277(49):47692 [9]Li J,Lu YM,Xue LX,et al.A structurally novel saltregulated promoter of duplicated carbonic anhydrase gene 1 from Dunaliella salina[J].Mol Biol Rep,2010,37(2):1143 [10]Fields S,Song O.A novel genetic system to detect proteinprotein interactions[J].Nature,1989,340(6230):245

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备注/Memo

备注/Memo:
*科技部国际科技合作基金资助项目2007DFA01240;国家自然科学基金资助项目30700014 #通讯作者,男,1944年2月生,教授,博士研究生导师,研究方向:肿瘤标志物与基因工程,Email:xuelx@zzu.edu.cn
更新日期/Last Update: 1900-01-01