Cao Jing, Huang Yu-Feng, Gao Jian, Wang Hao-Yang, Lu Jin-Chun
Department of Immunology, School of Preclinical Medicine, Nanjing Medical University, Nanjing, Jiangsu 210029, China.
Zhonghua Nan Ke Xue. 2009 Apr;15(4):318-21.
To construct a recombinant plasmid vector of the RV specific fragment for expressing the specific fragment of RV E1 protein.
RNA of the RV attenuated live vaccine Wistar RA27/3 strain was extracted and reversely transcribed. The specific fragment of the E1 gene was amplified and the PCR products cloned in the vector pGEX-2T after purification. Positive clones were selected and identified by two-enzyme digestion and sequence analysis.
A 330 bp target fragment was successfully cloned, and the sequence of the recombinant plasmid was consistent with the original sequence.
Successful cloning of the RV El specific fragment and the construction of the recombinant plasmid have laid a foundation for further expressing the recombinant protein.
构建用于表达轮状病毒(RV)E1蛋白特异性片段的RV特异性片段重组质粒载体。
提取RV减毒活疫苗Wistar RA27/3株的RNA并进行反转录。扩增E1基因的特异性片段,纯化后的PCR产物克隆至载体pGEX-2T中。通过双酶切和序列分析筛选并鉴定阳性克隆。
成功克隆出330 bp的目标片段,重组质粒序列与原始序列一致。
RV E1特异性片段的成功克隆及重组质粒的构建为进一步表达重组蛋白奠定了基础。