A hybrid plasmid pGEM-pET28 applied for heterologous
expression of Neospora caninum actin

Pereira, Luíz Miguel; Baroni, Luciana; Bezerra, Marcos Alexandre; Yatsuda, Ana Patrícia

doi:10.19185/matters.201702000012

Cited by 1 publication

(1 citation statement)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Under non-inducible conditions, the target gene can be silenced completely without transcription (89). Additionally, pET-28a(+) not only provides different tags (N-terminal: His/Thrombin/T7 protein tag and Cterminal: His tag) but also has appropriate restriction sites, which were beneficial to the purification of the target protein and cloning of different genes (90). However, Shilling etal.…”

Section: In Silico Molecular Cloningmentioning

confidence: 99%

Immunoinformatics Approach Toward the Introduction of a Novel Multi-Epitope Vaccine Against Clostridium difficile

et al. 2022

View full text Add to dashboard Cite

Clostridium difficile (C.difficile) is an exclusively anaerobic, spore-forming, and Gram-positive pathogen that is the most common cause of nosocomial diarrhea and is becoming increasingly prevalent in the community. Because C. difficile is strictly anaerobic, spores that can survive for months in the external environment contribute to the persistence and diffusion of C. difficile within the healthcare environment and community. Antimicrobial therapy disrupts the natural intestinal flora, allowing spores to develop into propagules that colonize the colon and produce toxins, thus leading to antibiotic-associated diarrhea and pseudomembranous enteritis. However, there is no licensed vaccine to prevent Clostridium difficile infection (CDI). In this study, a multi-epitope vaccine was designed using modern computer methods. Two target proteins, CdeC, affecting spore germination, and fliD, affecting propagule colonization, were chosen to construct the vaccine so that it could simultaneously induce the immune response against two different forms (spore and propagule) of C. difficile. We obtained the protein sequences from the National Center for Biotechnology Information (NCBI) database. After the layers of filtration, 5 cytotoxic T-cell lymphocyte (CTL) epitopes, 5 helper T lymphocyte (HTL) epitopes, and 7 B-cell linear epitopes were finally selected for vaccine construction. Then, to enhance the immunogenicity of the designed vaccine, an adjuvant was added to construct the vaccine. The Prabi and RaptorX servers were used to predict the vaccine’s two- and three-dimensional (3D) structures, respectively. Additionally, we refined and validated the structures of the vaccine construct. Molecular docking and molecular dynamics (MD) simulation were performed to check the interaction model of the vaccine–Toll-like receptor (TLR) complexes, vaccine–major histocompatibility complex (MHC) complexes, and vaccine–B-cell receptor (BCR) complex. Furthermore, immune stimulation, population coverage, and in silico molecular cloning were also conducted. The foregoing findings suggest that the final formulated vaccine is promising against the pathogen, but more researchers are needed to verify it.

show abstract

Section: In Silico Molecular Cloningmentioning

confidence: 99%