<abstract xmlns="http://www.w3.org/1999/xhtml">

<sec id="j_jvetres-2021-0011_s_005">
<h3>Introduction</h3>
<p>Ovine interferon-tau (oIFN-τ) is a newly discovered type I interferon. This study used biochemical techniques to transform the oIFN-τ gene into <italic>Escherichia coli</italic> to obtain the mass and soluble expression of the recombinant protein.</p>
</sec>
<sec id="j_jvetres-2021-0011_s_006">
<h3>Material and Methods</h3>
<p>First, total RNA was extracted from fresh sheep embryonic tissues with TRIzol reagent and then used as a template to reverse transcribe and amplify the mature oIFN-τ gene with RT-PCR. The amplified product was next digested with the <italic>Hind</italic>III and <italic>Xho</italic>I restriction enzymes and inserted into the pET-32a(+) vector to construct the prokaryotic expression plasmid. The corrected in-frame recombinant plasmid, pET-32a(+)-oIFN-τ, was transformed into <italic>E. coli</italic> Rosetta (DE3) competent cells. After induction with isopropyl-beta-D-thiogalactopyranoside (IPTG), the recombinant protein was detected in bacteria. Finally, the bacteria were lysed by sonication, and the recombinant protein was purified by nickel affinity chromatography and DEAE anion exchange chromatography.</p>
</sec>
<sec id="j_jvetres-2021-0011_s_007">
<h3>Results</h3>
<p>The protein was confirmed to be oIFN-τ, which mainly existed in the soluble lysate fraction, as proven by SDS-PAGE and Western blot assays.</p>
</sec>
<sec id="j_jvetres-2021-0011_s_008">
<h3>Conclusion</h3>
<p>Purified IFN-τ exists mostly in a soluble form, and its anti-vesicular stomatitis virus (VSV) activity reached 7.08×10(6)IU/mL.</p>
</sec>
</abstract>

Introduction
Ovine interferon-tau (oIFN-τ) is a newly discovered type I interferon. This study used biochemical techniques to transform the oIFN-τ gene into Escherichia coli to obtain the mass and soluble expression of the recombinant protein.


Material and Methods
First, total RNA was extracted from fresh sheep embryonic tissues with TRIzol reagent and then used as a template to reverse transcribe and amplify the mature oIFN-τ gene with RT-PCR. The amplified product was next digested with the HindIII and XhoI restriction enzymes and inserted into the pET-32a(+) vector to construct the prokaryotic expression plasmid. The corrected in-frame recombinant plasmid, pET-32a(+)-oIFN-τ, was transformed into E. coli Rosetta (DE3) competent cells. After induction with isopropyl-beta-D-thiogalactopyranoside (IPTG), the recombinant protein was detected in bacteria. Finally, the bacteria were lysed by sonication, and the recombinant protein was purified by nickel affinity chromatography and DEAE anion exchange chromatography.


Results
The protein was confirmed to be oIFN-τ, which mainly existed in the soluble lysate fraction, as proven by SDS-PAGE and Western blot assays.


Conclusion
Purified IFN-τ exists mostly in a soluble form, and its anti-vesicular stomatitis virus (VSV) activity reached 7.08×10(6)IU/mL.

Expression, purification, and bioactivity of a soluble recombinant ovine interferon-tau in <italic>Escherichia coli</italic>

Journal of Veterinary Research

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

Expression, purification, and bioactivity of a soluble recombinant ovine interferon-tau in 

Introduction
Ovine interferon-tau (oIFN-τ) is a newly discovered type I interferon. This study used biochemical techniques to transform the oIFN-τ gene into...

Expression, purification, and bioactivity of a soluble recombinant ovine interferon-tau in Escherichia coli

Data publikacji: 29 sty 2021

Zakres stron: 101 - 108

Otrzymano: 15 cze 2020

Przyjęty: 27 sty 2021

DOI: https://doi.org/10.2478/jvetres-2021-0011

Słowa kluczowe
ovine interferon-tau, cytopathic effect inhibition assay, soluble expression, purification, antiviral activity

© 2021 H.Y. Yu et al., published by Sciendo

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

Expression, purification, and bioactivity of a soluble recombinant ovine interferon-tau in Escherichia coli

Data publikacji: 29 sty 2021

Zakres stron: 101 - 108

Otrzymano: 15 cze 2020

Przyjęty: 27 sty 2021

DOI: https://doi.org/10.2478/jvetres-2021-0011

Słowa kluczoweovine interferon-tau, cytopathic effect inhibition assay, soluble expression, purification, antiviral activity

© 2021 H.Y. Yu et al., published by Sciendo

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

Słowa kluczowe
ovine interferon-tau, cytopathic effect inhibition assay, soluble expression, purification, antiviral activity