Expression of tomato yellow leaf curl virus coat protein using baculovirus expression system and evaluation of its utility as a viral antigen

Abstract

DNA encoding the coat protein (CP) of an Egyptian isolate of tomato yellow leaf curl virus (TYLCV) was inserted into the genome of Autographa californica nucleopolyhedrovirus (AcNPV) under the control of polyhedrin promoter. The generated recombinant baculovirus construct harboring the coat protein gene was characterized using PCR analysis. The recombinant coat protein expressed in infected insect cells was used as a coating antigen in an indirect Enzyme-linked immunosorbent assay (ELISA) and dot blot to test its utility for the detection of antibody generated against TYLCV virus particles. The results of ELISA and dot blot showed that the TYLCV-antibodies reacted positively with extracts of infected cells using the recombinant virus as a coating antigen with strong signals as well as the TYLCV infected tomato and beat plant extracts as positive samples. Scanning electron microscope examination showed that the expressed TYLCV coat protein was self-assembled into virus-like particles (VLPs) similar in size and morphology to TYLCV virus particles. These results concluded that, the expressed coat protein of TYLCV using baculovirus vector system is a reliable candidate for generation of anti-CP antibody for inexpensive detection of TYLCV-infected plants using indirect CP-ELISA or dot blot with high specificity.

Construction, Expression and Evaluation of Recombinant VP2 Protein for serotype-independent Detection of FMDV Seropositive Animals in Egypt

“…However, both techniques (LPBE and VNT) use inactivated live virus that poses certain health risks and they are also time-consuming, technically challenging, and have a significant rate of false positive reactions. Therefore, novel tests that harness the recombinant DNA technology to express target proteins in eukaryotic or prokaryotic systems are under development 11,12 .…”

Expression of 1B capsid protein of Foot-and-mouth disease virus (FMDV) using baculovirus expression system and its validation in detecting SAT 2- specific antisera

“…Reasons for using BacSF9 system for protein expression included (1) secretion into the growth medium; (2) production of less aggregated proteins with an increased probability of correct folding, and (3) post-translational modifications for better antigenicity and specificity. Previously, the baculovirus expression system was successfully used to express different viral proteins such as the tomato yellow leaf curl virus coat protein (TYLCV) (El-Gaied, Salem & Elmenofy, 2017), Garlic mite-borne filamentous virus (GarMbFV) (Ardisson-Arau'jo et al, 2013), and glycoprotein E (gE) of the Egyptian BoHV-1.1 Abu-Hammad strain (rBac/gE-AbuH) (El-Kholy et al, 2013;El-Gaied et al, 2020). Furthermore, recombinant proteins produced using this system were validated for their antigenicity.…”

Utilization of a recombinant defensin from Maize (Zea mays L.) as a potential antimicrobial peptide

“…The pGEX-4 T-1 harboring recombinant defensin (pGEX-4 T-1-Def) was transformed into the BL21 (DE3) E. coli strain (Elgaied et al 2017 ; Elmenofy et al 2020a ). Positive colonies were selected on the basis of their growth on Luria–Bertani (LB) agar plates supplemented with 50 µg/mL ampicillin.…”