Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus
IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (...
Ausführliche Beschreibung
Gespeichert in:
| Autor*in: |
Fu, Jianping [verfasserIn] |
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| Format: |
E-Artikel |
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| Sprache: |
Englisch |
| Erschienen: |
2019transfer abstract |
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| Schlagwörter: |
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| Umfang: |
4 |
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| Übergeordnetes Werk: |
Enthalten in: Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention - Istanbuly, Sedralmontaha ELSEVIER, 2021, London |
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| Übergeordnetes Werk: |
volume:88 ; year:2019 ; pages:403-406 ; extent:4 |
| Links: |
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| DOI / URN: |
10.1016/j.fsi.2019.03.022 |
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ELV046271368 |
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| 245 | 1 | 0 | |a Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus |
| 264 | 1 | |c 2019transfer abstract | |
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| 520 | |a IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. | ||
| 520 | |a IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. | ||
| 650 | 7 | |a Common carp |2 Elsevier | |
| 650 | 7 | |a SVCV |2 Elsevier | |
| 650 | 7 | |a IFN-γ |2 Elsevier | |
| 650 | 7 | |a Intein |2 Elsevier | |
| 700 | 1 | |a Yi, Zhiqiang |4 oth | |
| 700 | 1 | |a Cui, Hao |4 oth | |
| 700 | 1 | |a Song, Chunhui |4 oth | |
| 700 | 1 | |a Yu, Miao |4 oth | |
| 700 | 1 | |a Liu, Yi |4 oth | |
| 773 | 0 | 8 | |i Enthalten in |n Academic Press |a Istanbuly, Sedralmontaha ELSEVIER |t Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention |d 2021 |g London |w (DE-627)ELV006540406 |
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10.1016/j.fsi.2019.03.022 doi GBV00000000000584.pica (DE-627)ELV046271368 (ELSEVIER)S1050-4648(19)30170-6 DE-627 ger DE-627 rakwb eng 610 VZ 44.85 bkl Fu, Jianping verfasserin aut Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus 2019transfer abstract 4 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. Common carp Elsevier SVCV Elsevier IFN-γ Elsevier Intein Elsevier Yi, Zhiqiang oth Cui, Hao oth Song, Chunhui oth Yu, Miao oth Liu, Yi oth Enthalten in Academic Press Istanbuly, Sedralmontaha ELSEVIER Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention 2021 London (DE-627)ELV006540406 volume:88 year:2019 pages:403-406 extent:4 https://doi.org/10.1016/j.fsi.2019.03.022 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U SSG-OLC-PHA 44.85 Kardiologie Angiologie VZ AR 88 2019 403-406 4 |
| spelling |
10.1016/j.fsi.2019.03.022 doi GBV00000000000584.pica (DE-627)ELV046271368 (ELSEVIER)S1050-4648(19)30170-6 DE-627 ger DE-627 rakwb eng 610 VZ 44.85 bkl Fu, Jianping verfasserin aut Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus 2019transfer abstract 4 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. Common carp Elsevier SVCV Elsevier IFN-γ Elsevier Intein Elsevier Yi, Zhiqiang oth Cui, Hao oth Song, Chunhui oth Yu, Miao oth Liu, Yi oth Enthalten in Academic Press Istanbuly, Sedralmontaha ELSEVIER Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention 2021 London (DE-627)ELV006540406 volume:88 year:2019 pages:403-406 extent:4 https://doi.org/10.1016/j.fsi.2019.03.022 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U SSG-OLC-PHA 44.85 Kardiologie Angiologie VZ AR 88 2019 403-406 4 |
| allfields_unstemmed |
10.1016/j.fsi.2019.03.022 doi GBV00000000000584.pica (DE-627)ELV046271368 (ELSEVIER)S1050-4648(19)30170-6 DE-627 ger DE-627 rakwb eng 610 VZ 44.85 bkl Fu, Jianping verfasserin aut Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus 2019transfer abstract 4 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. Common carp Elsevier SVCV Elsevier IFN-γ Elsevier Intein Elsevier Yi, Zhiqiang oth Cui, Hao oth Song, Chunhui oth Yu, Miao oth Liu, Yi oth Enthalten in Academic Press Istanbuly, Sedralmontaha ELSEVIER Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention 2021 London (DE-627)ELV006540406 volume:88 year:2019 pages:403-406 extent:4 https://doi.org/10.1016/j.fsi.2019.03.022 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U SSG-OLC-PHA 44.85 Kardiologie Angiologie VZ AR 88 2019 403-406 4 |
| allfieldsGer |
10.1016/j.fsi.2019.03.022 doi GBV00000000000584.pica (DE-627)ELV046271368 (ELSEVIER)S1050-4648(19)30170-6 DE-627 ger DE-627 rakwb eng 610 VZ 44.85 bkl Fu, Jianping verfasserin aut Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus 2019transfer abstract 4 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. Common carp Elsevier SVCV Elsevier IFN-γ Elsevier Intein Elsevier Yi, Zhiqiang oth Cui, Hao oth Song, Chunhui oth Yu, Miao oth Liu, Yi oth Enthalten in Academic Press Istanbuly, Sedralmontaha ELSEVIER Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention 2021 London (DE-627)ELV006540406 volume:88 year:2019 pages:403-406 extent:4 https://doi.org/10.1016/j.fsi.2019.03.022 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U SSG-OLC-PHA 44.85 Kardiologie Angiologie VZ AR 88 2019 403-406 4 |
| allfieldsSound |
10.1016/j.fsi.2019.03.022 doi GBV00000000000584.pica (DE-627)ELV046271368 (ELSEVIER)S1050-4648(19)30170-6 DE-627 ger DE-627 rakwb eng 610 VZ 44.85 bkl Fu, Jianping verfasserin aut Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus 2019transfer abstract 4 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. Common carp Elsevier SVCV Elsevier IFN-γ Elsevier Intein Elsevier Yi, Zhiqiang oth Cui, Hao oth Song, Chunhui oth Yu, Miao oth Liu, Yi oth Enthalten in Academic Press Istanbuly, Sedralmontaha ELSEVIER Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention 2021 London (DE-627)ELV006540406 volume:88 year:2019 pages:403-406 extent:4 https://doi.org/10.1016/j.fsi.2019.03.022 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U SSG-OLC-PHA 44.85 Kardiologie Angiologie VZ AR 88 2019 403-406 4 |
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Comparison of Outcomes of Patients With Versus Without Chronic Liver Disease Undergoing Percutaneous Coronary Intervention |
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intein-mediated expression and purification of common carp ifn-γ and its protective effect against spring viremia of carp virus |
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Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus |
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IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. |
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IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. |
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IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture. |
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Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus |
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