OpenLink Software

About: AIM: Generation of recombinant human adenovirus type 5 expressing foot-and-mouth disease virus (FMDV) capsid protein genes along with full-length 2B, 3B and 3C(pro) and its characterization. MATERIALS AND METHODS: FMD viral RNA isolation, cDNA synthesis, and polymerase chain reaction were performed to synthesize expression cassettes (P1-2AB3BC(wt) and P1-2AB3BC(m)) followed by cloning in pShuttle-CMV vector. Chemically competent BJ5183-AD-1 cells were transformed with the recombinant pShuttle-CMV to produce recombinant adenoviral plasmids. HEK-293 cells were transfected with the recombinant adenoviral plasmids to generate recombinant adenoviruses (hAd5/P1-2AB3BC(wt) and hAd5/P1-2AB3BC(m)). Expression of the target proteins was analyzed by sandwich ELISA and indirect immunofluorescence assay. The recombinant adenoviruses were purified and concentrated by CsCl density gradient ultracentrifugation. Growth kinetics and thermostability of the recombinant adenoviruses were compared with that of non-recombinant replication-defective adenovirus (dAd5). RESULTS: The recombinant adenoviruses containing capsid protein genes of the FMDV O/IND/R2/75 were generated and amplified in HEK-293 cells. The titer of the recombinant adenoviruses was approximately 10(8), 10(9.5) and 10(11) TCID(50)/ml in supernatant media, cell lysate and CsCl purified preparation, respectively. Expression of the FMDV capsid protein was detectable in sandwich ELISA and confirmed by immunofluorescence assay. Growth kinetics of the recombinant adenoviruses did not reveal a significant difference when compared with that of dAd5. A decrement of up to 10-fold at 4°C and 21-fold at 37°C was recorded in the virus titers during 60 h incubation period and found to be statistically significant (p<0.01). CONCLUSION: Recombinant adenoviruses expressing capsid proteins of the FMDV O/IND/R2/75 were constructed and produced in high titers. In vitro expression of the target proteins in the adenovirus vector system was detected by sandwich ELISA and immunofluorescence assay.

 Permalink

an Entity references as follows:

Subject of Sentences In Document
Object of Sentences In Document
Explicit Coreferences
Implicit Coreferences
Faceted Search & Find service v1.13.91

Alternative Linked Data Documents: Sponger | ODE     Raw Data in: CXML | CSV | RDF ( N-Triples N3/Turtle JSON XML ) | OData ( Atom JSON ) | Microdata ( JSON HTML) | JSON-LD    About   
This material is Open Knowledge   W3C Semantic Web Technology [RDF Data] This material is Open Knowledge Creative Commons License Valid XHTML + RDFa
This work is licensed under a Creative Commons Attribution-Share Alike 3.0 Unported License.
OpenLink Virtuoso version 07.20.3229 as of Jul 10 2020, on Linux (x86_64-pc-linux-gnu), Single-Server Edition (94 GB total memory)
Copyright © 2009-2025 OpenLink Software