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Updated: Sep 20, 2026
Simplified Reverse Genetics Method to Recover Recombinant Rotaviruses Expressing Reporter Proteins
Published on: April 17, 2020
[Expression and glycosylation of rotavirus strain SA11 VP4 protein in a recombinant adenovirus]
1Department of Molecular Biology, Institute of Medical Biology, CAMS, PUMC, Kunming 650107, China.
Objective:
A modified VP4 gene of rotavirus SA11 strain was expressed by a recombinant human adenovirus.
Methods:
A whole VP4 gene was obtained with PCR and induced the signal peptide at the gene N terminal. The chimera gene was cloned into pCMV plasmid that consist of human cytomagolovirus promoter and then cloned the gene to transfer human adenovirus type 5 vector. Homologues recombinant was performed by co-transfection to 293 cell line with recombinant plasmid and viral genome using CaPO4 precipitation.
Results:
VP4 gene is 2,362 base pair in length mutation was not found in whole VP4 gene sequence. Expressed product in recombinant adnovirus was confirmed to be specific and more antigenicity by indirect immunofluorescence assay. Both Western blot and immunoprecipitation assays showed that expressed protein molecular weight was higher than wild type VP4 protein and that modified product was corresponding to a glycosylation of VP4 protein.
Conclusions:
It may be a effective method to modify interested gene for enhancing stability, antigenicity and immunogenicity of expressive product.
