Luciferase:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Reporter Assay:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Virus:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Western Blot:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Real-time Polymerase Chain Reaction:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Immunofluorescence:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Activity Assay:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Construct:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
Plasmid Preparation:Article Title: Black carp IKKε collaborates with IRF3 in the antiviral signaling.
Article Snippet: Polyethylenimine (PEI) (polyscience, USA) was used for the transfection of HEK293T and EPC cells; Lipomax (SUDGEN, China) was used for MPK cells [23]. .. CIK cells were used to produce GCRV; EPC cells were used for produce SVCV, dual luciferase reporter assay, virus titers test and immunoblot assay; MPK cells were used for quantitative real-time PCR (qPCR) and immunofluorescence. pcDNA5/FRT/TO-HA, pcDNA5/FRT/TO-Flag, pEGFP-N1, pRL-TK, Luci-bcIFNa (for black carp IFNa promoter activity analysis), LuciDrIFNφ1pro (for zebrafish IFNφ1 promoter activity analysis), pLKO.1TRC (Addgene, USA) and pcDNA5/FRT/TO-Flag-bcIKKε were kept in the lab [24]. pcDNA5/FRT/TO-HA-bcIRF3 was constructed by inserting the open reading frame (ORF) of bcIRF3 into pcDNA5/FRT/TO between Kpn I and Xho I restriction sites with a HA tag at its N-terminus. bcIRF3-pEGFP-N1 was constructed by inserting the ORF of bcIRF3 into upstream of the pEGFP-N1 vector enhanced green fluorescent protein. .. The shRNA primers targeting the CDS of bcIRF3 gene were designed according to the company’s website (http://rnaidesigner.thermofisher. com/rnaiexpress) and were sub-cloned into pLKO.1-TRC after annealed as previously [24].
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