| 大菱鲆(Scophthalmus maximus)蛋白质二硫键异构酶SmPDIA3的表达分析和功能验证 |
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| 引用本文： | 唐启政,孙志宾,王新安,马爱军,杨双双,黄智慧. 大菱鲆(Scophthalmus maximus)蛋白质二硫键异构酶SmPDIA3的表达分析和功能验证[J]. 海洋与湖沼, 2019, 50(2): 409-419 |
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| 作者姓名： | 唐启政 孙志宾 王新安 马爱军 杨双双 黄智慧 |
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| 作者单位： | 中国水产科学研究院黄海水产研究所 山东省海洋渔业生物技术与遗传育种重点实验室 青岛市海水鱼类种子工程与生物技术重点实验室 青岛 266071;上海海洋大学水产与生命学院 上海201306;青岛海洋科学与技术国家实验室海洋生物学与生物技术功能实验室 青岛 266071;中国水产科学研究院黄海水产研究所 山东省海洋渔业生物技术与遗传育种重点实验室 青岛市海水鱼类种子工程与生物技术重点实验室 青岛 266071;青岛海洋科学与技术国家实验室海洋生物学与生物技术功能实验室 青岛 266071 |
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| 基金项目： | 现代农业产业技术体系专项，CARS-47-G01号；青岛海洋科学与技术国家实验室“鳌山人才”培养计划项目，2017ASTCP-OS04号；国家自然科学基金项目，41706168号；山东省重点研发计划，2016GSF115019号；山东省良种工程，2016LZGC031号。 |
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| 摘 要： | 本研究利用SMART-RACE技术克隆了大菱鲆SmPDIA3基因。SmPDIA3基因cDNA序列全长2083bp，包括1479bp的开放阅读框，编码492个氨基酸，GenBank登录号：MG765516。组织表达分析发现：SmPDIA3基因在肠、鳃、肝脏等组织中均有表达，其中在肝脏中表达量最高，脑中最低。进一步研究了温度胁迫后SmPDIA3基因在肠和肝脏组织中的表达变化规律，发现随着胁迫温度的升高，SmPDIA3基因的相对表达量总体上升，并在28℃时达到最大。利用原核表达技术，构建了pET-28a-PDIA3原核表达载体，IPTG诱导后融合蛋白主要在上清中表达。将上清中的蛋白分离纯化后，利用Western blot技术验证了目的蛋白的准确性。将纯化后的目的蛋白浓缩后，利用BCA蛋白浓度测定试剂盒测得蛋白浓度为243.18μg/mL。最后设计变性溶菌酶的复性实验，验证了SmPDIA3蛋白具有分子伴侣功能，能够辅助变性蛋白的正确折叠。
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| 关 键 词： | 大菱鲆 RACE SmPDIA3 原核表达 分子伴侣 |
| 收稿时间： | 2018-10-22 |
| 修稿时间： | 2018-12-06 |
EXPRESSION ANALYSIS AND FUNCTIONAL VERIFICATION OF PROTEIN DISULFIDE ISOMERASE SmPDIA3 IN TURBOT SCOPHTHALMUS MAXIMUS |
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| TANG Qi-Zheng,SUN Zhi-Bin,WANG Xin-An,MA Ai-Jun,YANG Shuang-Shuang and HUANG Zhi-Hui. EXPRESSION ANALYSIS AND FUNCTIONAL VERIFICATION OF PROTEIN DISULFIDE ISOMERASE SmPDIA3 IN TURBOT SCOPHTHALMUS MAXIMUS[J]. Oceanologia Et Limnologia Sinica, 2019, 50(2): 409-419 |
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| Authors: | TANG Qi-Zheng SUN Zhi-Bin WANG Xin-An MA Ai-Jun YANG Shuang-Shuang HUANG Zhi-Hui |
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| Affiliation: | Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Shandong Key Laboratory of Marine Fisheries Biotechnology and Genetic Breeding;Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Qingdao 266071, China;College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China;Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266071, China,Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Shandong Key Laboratory of Marine Fisheries Biotechnology and Genetic Breeding;Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Qingdao 266071, China;Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266071, China,Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Shandong Key Laboratory of Marine Fisheries Biotechnology and Genetic Breeding;Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Qingdao 266071, China;Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266071, China,Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Shandong Key Laboratory of Marine Fisheries Biotechnology and Genetic Breeding;Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Qingdao 266071, China;Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266071, China,Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Shandong Key Laboratory of Marine Fisheries Biotechnology and Genetic Breeding;Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Qingdao 266071, China;Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266071, China and Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences;Shandong Key Laboratory of Marine Fisheries Biotechnology and Genetic Breeding;Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Qingdao 266071, China;Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266071, China |
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| Abstract: | |
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| Keywords: | turbot Scophthalmus maximus RACE SmPDIA3 prokaryotic expression molecular chaperone |
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