Search citation statements
Paper Sections
Citation Types
Year Published
Publication Types
Relationship
Authors
Journals
Porcine circovirus-associated diseases (PCVDs) are among the most significant challenges for pig farming in developed countries. Porcine circovirus type 2 (PCV-2) is considered the main etiological agent of postweaning multisystemic wasting syndrome in piglets. Mass PCVD occurrence has been reported in most regions of the world, that results in serious economic consequences. Optimal PCVD prevention is known to be achieved through a set of veterinary and sanitary measures in combination with vaccination. High evolutionary virus variability facilitating new genotype and strain emergence requires development of new candidate recombinant vaccines against PCV-2 infection. The study was aimed at construction of prokaryotic system for PCV-2 ORF-2 gene fragment expression and its functionality assessment. A genetic insert constructed from the most immunogenic type-specific PCV-2 epitopes based on genotype 2a, 2b, 2d strain and isolate consensus sequence was cloned into the expression vector pET-22b(+) that was incorporated into the Escherichia coli strain Rosetta 2(DE3). The transformants were selected based on the marker gene of ampicillin resistance on a selective medium. Target gene expression was induced by adding of isopropyl-β-D-1-thiogalactopyranoside at different concentrations. As a result, Escherichia coli Rosetta 2(DE3)/pET-22b-ORF-2 strain, a producer of capsid protein fragment (92–233 amino acid residues), was constructed. It was found that in the presence of 1 mM isopropyl-β-D-1-thiogalactopyranoside, the expression level of soluble truncated rCap was 35–40 mg/L 6 hours after induction. The expression product was tested for its specificity with indirect ELISA using whole-virion PCV-2-hyperimmunized porcine serum. It was shown that the positivity coefficient of producer strain cell lysates averaged to 4.34 (p < 0.005). The recombinant rCap protein is suitable for serological diagnosis and is also of interest as a vaccine component, which is the goal of our further studies.
Porcine circovirus-associated diseases (PCVDs) are among the most significant challenges for pig farming in developed countries. Porcine circovirus type 2 (PCV-2) is considered the main etiological agent of postweaning multisystemic wasting syndrome in piglets. Mass PCVD occurrence has been reported in most regions of the world, that results in serious economic consequences. Optimal PCVD prevention is known to be achieved through a set of veterinary and sanitary measures in combination with vaccination. High evolutionary virus variability facilitating new genotype and strain emergence requires development of new candidate recombinant vaccines against PCV-2 infection. The study was aimed at construction of prokaryotic system for PCV-2 ORF-2 gene fragment expression and its functionality assessment. A genetic insert constructed from the most immunogenic type-specific PCV-2 epitopes based on genotype 2a, 2b, 2d strain and isolate consensus sequence was cloned into the expression vector pET-22b(+) that was incorporated into the Escherichia coli strain Rosetta 2(DE3). The transformants were selected based on the marker gene of ampicillin resistance on a selective medium. Target gene expression was induced by adding of isopropyl-β-D-1-thiogalactopyranoside at different concentrations. As a result, Escherichia coli Rosetta 2(DE3)/pET-22b-ORF-2 strain, a producer of capsid protein fragment (92–233 amino acid residues), was constructed. It was found that in the presence of 1 mM isopropyl-β-D-1-thiogalactopyranoside, the expression level of soluble truncated rCap was 35–40 mg/L 6 hours after induction. The expression product was tested for its specificity with indirect ELISA using whole-virion PCV-2-hyperimmunized porcine serum. It was shown that the positivity coefficient of producer strain cell lysates averaged to 4.34 (p < 0.005). The recombinant rCap protein is suitable for serological diagnosis and is also of interest as a vaccine component, which is the goal of our further studies.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.