Effects of genomic location of foreign gene in measles virus vector on gene expression and virus replication
10.3760/cma.j.cn112309-20220329-00091
- VernacularTitle:麻疹病毒载体中外源基因插入位置对基因表达及病毒复制的影响
- Author:
Wenhao SU
1
;
Xiuxiu REN
;
Tingting ZHAO
;
Yinan WANG
;
Shishi LI
;
Qiufang HUANG
;
Xiaojie WANG
;
Xiaohuan ZHANG
;
Jiangbo WEI
Author Information
1. 国药中生生物技术研究院卫江波实验室，北京 101111
- Keywords:
Measles virus Shanghai-191 strain;
Measles virus vector;
Reverse genetics;
S1 protein
- From:
Chinese Journal of Microbiology and Immunology
2022;42(12):924-930
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effects of genomic location of a foreign gene in Shanghai-191 strain measles virus (MV) vector on gene expression and virus replication.Methods:The nucleotide sequence encoding S1 protein of SARS-CoV-2 was inserted at different positions in MV antigenome (the upstream of the N gene, between P and M genes, between H and L genes), and co-transfected into 293T cells with helper plasmids coding T7 RNA polymerase and N, P, and L proteins, respectively. The transfected cells were lysed and the supernatants were used to infected Vero cells to harvest recombinant viruses. S1 proteins expressed by the recombinant viruses were identified by RT-PCR, indirect immunofluorescence assay, Western blot and ELISA. Growth kinetics of the recombinant viruses were analyzed.Results:Recombinant viruses were failed to be rescued when the S1 protein-coding sequence was cloned into the upstream of N gene. Two recombinant viruses, MV-M-S1 and MV-L-S1, were successfully rescued when cloning the S1 protein-coding sequence into the intergenic region between P and M genes, or H and L genes, and could express S1 protein. MV-M-S1 expressed more S1 protein than MV-L-S1, but the titer of MV-M-S1 was lower.Conclusions:Inserting a foreign gene at different positions in the MV genome might have different effects on gene expression and virus replication. This study provided reference for the subsequent construction of MV vector.
