<abstract xmlns="http://www.w3.org/1999/xhtml"><bold>Introduction:</bold> The development of Jembrana disease vaccine is an important effort to prevent losses in the Bali cattle industry in Indonesia. This study aims to prepare a Jembrana DNA vaccine encoding the transmembrane portion of the envelope protein in pEGFP-C1 and test the success of its delivery in culture cells using a chitosan-DNA complex.<bold>Material and Methods:</bold> Cloning of the DNA vaccine was successfully performed on <italic>E. coli</italic> DH5α and confirmed by colony PCR, restriction analysis and sequencing. The plasmids were prepared as a chitosan complex using the complex coacervation method and physicochemically characterised using a particle size analyser. A transfection assay was performed in HeLa cells with 4 h exposure, and mRNA expression was assessed at 24 h post transfection.<bold>Results:</bold> With a 1:2 (wt./wt.) ratio of DNA and chitosan, the complexes have a mean diameter of 236 nm, zeta potential value of + 17.9 mV, and showed no high toxicity potential in the HeLa cells. This complex successfully delivered the DNA into cells, as shown by the presence of a specific RT-PCR product (336 bp). However, the real-time PCR analysis showed that the delivery with chitosan complex resulted in lower target mRNA expression when compared with a commercial transfecting agent.<bold>Conclusion:</bold> pEGFP-env-tm JDV as a candidate vaccine can be delivered as the chitosan-DNA complex and be expressed at the transcription level <italic>in vitro</italic>. This initial study will be used for further improvement and evaluation <italic>in vivo</italic>.</abstract>

Introduction: The development of Jembrana disease vaccine is an important effort to prevent losses in the Bali cattle industry in Indonesia. This study aims to prepare a Jembrana DNA vaccine encoding the transmembrane portion of the envelope protein in pEGFP-C1 and test the success of its delivery in culture cells using a chitosan-DNA complex.Material and Methods: Cloning of the DNA vaccine was successfully performed on E. coli DH5α and confirmed by colony PCR, restriction analysis and sequencing. The plasmids were prepared as a chitosan complex using the complex coacervation method and physicochemically characterised using a particle size analyser. A transfection assay was performed in HeLa cells with 4 h exposure, and mRNA expression was assessed at 24 h post transfection.Results: With a 1:2 (wt./wt.) ratio of DNA and chitosan, the complexes have a mean diameter of 236 nm, zeta potential value of + 17.9 mV, and showed no high toxicity potential in the HeLa cells. This complex successfully delivered the DNA into cells, as shown by the presence of a specific RT-PCR product (336 bp). However, the real-time PCR analysis showed that the delivery with chitosan complex resulted in lower target mRNA expression when compared with a commercial transfecting agent.Conclusion: pEGFP-env-tm JDV as a candidate vaccine can be delivered as the chitosan-DNA complex and be expressed at the transcription level in vitro. This initial study will be used for further improvement and evaluation in vivo.

<italic>In vitro</italic> evaluation of chitosan-DNA plasmid complex encoding Jembrana disease virus Env-TM protein as a vaccine candidate

Department of Reproduction and Obstetrics, Faculty of Veterinary Medicine

Journal of Veterinary Research

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

 evaluation of chitosan-DNA plasmid complex encoding Jembrana disease virus Env-TM protein as a vaccine candidate

Introduction: The development of Jembrana disease vaccine is an important effort to prevent losses in the Bali cattle industry in Indonesia. This study aims to...

In vitro evaluation of chitosan-DNA plasmid complex encoding Jembrana disease virus Env-TM protein as a vaccine candidate

Pubblicato online: 28 mar 2019

Pagine: 7 - 16

Ricevuto: 27 set 2018

Accettato: 01 mar 2019

DOI: https://doi.org/10.2478/jvetres-2019-0018

Parole chiave
chitosan-DNA complex, gene, Jembrana disease virus, pEGFP-C1, vaccine

© 2019 Januar Ishak et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

In vitro evaluation of chitosan-DNA plasmid complex encoding Jembrana disease virus Env-TM protein as a vaccine candidate

Pubblicato online: 28 mar 2019

Pagine: 7 - 16

Ricevuto: 27 set 2018

Accettato: 01 mar 2019

DOI: https://doi.org/10.2478/jvetres-2019-0018

Parole chiavechitosan-DNA complex, gene, Jembrana disease virus, pEGFP-C1, vaccine

© 2019 Januar Ishak et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Parole chiave
chitosan-DNA complex, gene, Jembrana disease virus, pEGFP-C1, vaccine