Abstract:Objective: To construct the recombinant lentivirus carrying the intracellular Notch1(ICN), and to validate its role in Notch signaling pathway and the proliferative capacity of Kaposi′s sarcomaassociated herpesvirus(KSHV)infected primary effusion lymphoma (PEL) cells. Methods: The sequences of ICN were cloned from expression plasmid pIPFlagICN and inserted into lentiviral plasmid pHAGECMVMCSIzsGreen(pHAGE) to construct the recombinant plasmid pHAGEICN, after which it was cotransfected with the package plasmid psPAX2 and the envelope plasmid pMD2.G into 293 T cells. The titer of the recombinant lentivirus expressing ICN was determined by gradient dilution. The recombinant lentivirus was transduced into BCP1 cells followed by detecting the expressions of ICN and Notch downstream effector genes by Western blotting and examining the proliferation of BCP1 cells by cell counting. Results: The results of enzyme digestion and DNA sequencing indicated that the recombinant plasmid pHAGEICN was successfully constructed. The titer of the recombinant lentivirus was about 2×107 TU/mL. By Western blotting, activated Notch signaling was found in ICN overexpressed BCP1 cells.The result from cell counting showed that overexpression of ICN could promote the proliferation of BCP1 cells. Conclusion: Recombinant lentiviral vector containing ICN has been successfully constructed, which can activate Notch signaling pathway and regulate the proliferation of PEL cells.
收稿日期: 2018-01-23
通讯作者:
卢春(通讯作者),教授,博士生导师,E-mail:clu@njmu.edu.cn
作者简介: 赵润然(1994—),女,硕士研究生
引用本文:
赵润然, 沈忱悠, 严沁, 卢春. Notch1蛋白胞内段重组慢病毒载体的构建及其在原发性渗出性淋巴瘤细胞中的功能验证[J]. 江苏大学学报:医学版, 2018, 28(02): 121-125.
ZHAO Run-ran, SHEN Chen-you, YAN Qin, LU Chun. Construction of recombinant expression lentivirus vector carrying intracellular Notch1 and its function validation in primary effusion lymphoma cells
. Journal of Jiangsu University(Medicine Edition), 2018, 28(02): 121-125.
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