| 小麦蓝矮植原体免疫膜蛋白(Imp)的跨膜区影响其在大肠杆菌中的表达 |
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| 引用本文： | 孙润红,于祥泉,陶烨,杨洋,吴云锋,张银武,李艳.小麦蓝矮植原体免疫膜蛋白(Imp)的跨膜区影响其在大肠杆菌中的表达[J].植物病理学报,2010,40(6):587-592. |
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| 作者姓名： | 孙润红 于祥泉 陶烨 杨洋 吴云锋 张银武 李艳 |
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| 作者单位： | 1 西北农林科技大学植物保护学院, 陕两省农业分子生物学重点实验宦, 杨凌 712100;2 西北农林科技大学生命科学学院, 杨凌 712100;3 河南驻马店上蔡县农技站, 上蔡 463800 |
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| 基金项目： | 国家自然科学基金，*，高等学校学科创新引智计划 |
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| 摘 要： | 根据跨膜区预测结果,设计ImpF1/ImpR1和ImpF2/ImpR2两对特异性引物。以ImpF1/ImpR1,ImpF1/ImpR2,Imp-F2/ImpR1和ImpF2/ImpR2共4个组合经PCR扩增得到4个目标片段,即:免疫膜蛋白Imp基因的全长(Imp-B);切除C-端跨膜区的Imp基因的基因序列(Imp-N);切除N-端跨膜区的Imp基因的基因序列(Imp-C);切除N-和C-端跨膜区的Imp基因的基因序列(Imp-S)。经酶切连接到原核表达载体Pmal-c2x,并转入E.coliBL21(DE3)PlysS菌株中。SDS-PAGE电泳验证,只有切除N-、C-端跨膜区的Imp基因的基因序列(Imp-S)得到大量表达,结果表明:小麦蓝矮植原体免疫膜蛋白的跨膜区影响其在大肠杆菌中的表达。根据跨膜区预测结果,设计ImpF1/ImpR1和ImpF2/ImpR2两对特异性引物。以ImpF1/ImpR1,ImpF1/ImpR2,Imp-F2/ImpR1和ImpF2/ImpR2共4个组合经PCR扩增得到4个目标片段,即:免疫膜蛋白Imp基因的全长(Imp-B);切除C-端跨膜区的Imp基因的基因序列(Imp-N);切除N-端跨膜区的Imp基因的基因序列(Imp-C);切除N-和C-端跨膜区的Imp基因的基因序列(Imp-S)。经酶切连接到原核表达载体Pmal-c2x,并转入E.coliBL21(DE3)PlysS菌株中。SDS-PAGE电泳验证,只有切除N-、C-端跨膜区的Imp基因的基因序列(Imp-S)得到大量表达,结果表明:小麦蓝矮植原体免疫膜蛋白的跨膜区影响其在大肠杆菌中的表达。
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| 关 键 词： | 小麦蓝矮植原表体 免疫膜蛋白 跨膜区 |
Influence of the transmembrane region of immunodominant membrane protein (Imp) from wheat blue dwarf phytoplasma on Imp expression in Escherichia coli |
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| SUN Run-Hong,YU Xiang-Quan,TAO Ye,YANG Yang,WU Yun-Feng,ZHANG Yin-Wu,LI Yan.Influence of the transmembrane region of immunodominant membrane protein (Imp) from wheat blue dwarf phytoplasma on Imp expression in Escherichia coli[J].Acta Phytopathologica Sinica,2010,40(6):587-592. |
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| Authors: | SUN Run-Hong YU Xiang-Quan TAO Ye YANG Yang WU Yun-Feng ZHANG Yin-Wu LI Yan |
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| Institution: | 1 College of Plant Protection, Northwest A & F University, Shaanxi Key Laboratory of Molecular Biology for Agriculture, Yangling 712100, China;2 Colleges of Life Sciences, Northwest A & F University, Yangling 712100, China;3 Henan Shangcai Agricultural Technology Service Center, Shangcai 463800, China |
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| Abstract: | Two pairs of specific primers, ImpF1/ImpR1 and ImpF2/ImpR2, were designed according to the prediction result of possible transmembrane helices for immunodominant membrane protein(Imp) of wheat blue dwarf(WBD) phytoplasma.Four sequences, Imp-B, Imp-N, Imp-C and Imp-S were respectively obtained by using combined primers ImpF1/ImpR1, ImpF1/ImpR2, ImpF2/ImpR1 and ImpF2/ImpR2.The overall length of Imp was named Imp-B;the Imp sequence excision of the C-transmembrane region was Imp-N;the Imp excision of the N-transmembrane region was Imp-C and the Imp excision of the C-and N-transmembrane regions was Imp-S.After double enzyme digestion, these four sequences were respectively sub-cloned into the expression vector Pmal-c2x and then transformed into E.coli strain BL21(DE3) PlysS.SDS-PAGE analysis showed that only Imp-S with cleaved N-terminal and C-terminal transmembrane regions could be expressed.The results demonstrated that the transmembrane region can influence the Imp expression of WBD phytoplasma in E.coli. |
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| Keywords: | wheat blue dwarf phytoplasma immunodominant membrane protein(Imp) transmembrane region |
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