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    GE Healthcare pgex 4t 1
    Cloning strategies for cloning the paracoccin ORF for expression. The left panel shows the standard strategy, with PCR amplification of the largest exon, restriction endonuclease digestion, and cloning into the expression vector <t>pGEX-4T-1.</t> The positions of the exons are displayed on the map of the gene. Genomic DNA template was extracted from P. brasiliensis strain Pb18. Agarose gel electrophoresis (mid-left) shows the corresponding band amplified by PCR. The exon 4 amplicon was cloned by Bam HI and Eco RI digestion. The right panel shows the strategy for synthesis of the predicted paracoccin sequence fused with the 5′-UTR elements for transcription in the vector pUC57. Green arrow, T7 promoter; black box, lacO (lac operator); and red box, the phage T7 trailer sequence for ribosome binding.
    Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 96/100, based on 3798 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare xhoi digested pgex 4t 1
    Cloning strategies for cloning the paracoccin ORF for expression. The left panel shows the standard strategy, with PCR amplification of the largest exon, restriction endonuclease digestion, and cloning into the expression vector <t>pGEX-4T-1.</t> The positions of the exons are displayed on the map of the gene. Genomic DNA template was extracted from P. brasiliensis strain Pb18. Agarose gel electrophoresis (mid-left) shows the corresponding band amplified by PCR. The exon 4 amplicon was cloned by Bam HI and Eco RI digestion. The right panel shows the strategy for synthesis of the predicted paracoccin sequence fused with the 5′-UTR elements for transcription in the vector pUC57. Green arrow, T7 promoter; black box, lacO (lac operator); and red box, the phage T7 trailer sequence for ribosome binding.
    Xhoi Digested Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare gst aurora bwt
    Cloning strategies for cloning the paracoccin ORF for expression. The left panel shows the standard strategy, with PCR amplification of the largest exon, restriction endonuclease digestion, and cloning into the expression vector <t>pGEX-4T-1.</t> The positions of the exons are displayed on the map of the gene. Genomic DNA template was extracted from P. brasiliensis strain Pb18. Agarose gel electrophoresis (mid-left) shows the corresponding band amplified by PCR. The exon 4 amplicon was cloned by Bam HI and Eco RI digestion. The right panel shows the strategy for synthesis of the predicted paracoccin sequence fused with the 5′-UTR elements for transcription in the vector pUC57. Green arrow, T7 promoter; black box, lacO (lac operator); and red box, the phage T7 trailer sequence for ribosome binding.
    Gst Aurora Bwt, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare gst pgex 4t 1
    Cloning strategies for cloning the paracoccin ORF for expression. The left panel shows the standard strategy, with PCR amplification of the largest exon, restriction endonuclease digestion, and cloning into the expression vector <t>pGEX-4T-1.</t> The positions of the exons are displayed on the map of the gene. Genomic DNA template was extracted from P. brasiliensis strain Pb18. Agarose gel electrophoresis (mid-left) shows the corresponding band amplified by PCR. The exon 4 amplicon was cloned by Bam HI and Eco RI digestion. The right panel shows the strategy for synthesis of the predicted paracoccin sequence fused with the 5′-UTR elements for transcription in the vector pUC57. Green arrow, T7 promoter; black box, lacO (lac operator); and red box, the phage T7 trailer sequence for ribosome binding.
    Gst Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 86/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    GE Healthcare plasmid pgex 4t 1
    VP1 protein expression in E. coli using various expression vectors . The VP1 protein expression in E. coli BL21(DE3) using the expression vectors, pET28a (A) and <t>pGEX-4T-1</t> (B) was analyzed by SDS-PAGE and Western-blotting. Anti-His tag and anti-GST tag monoclonal antibody were respectively used for the recognition of differently tagged VP1 proteins expressed by the different expression vectors used. Lane M, pre-stained protein marker; lane 1 and lane 3, before IPTG induction; lane 2 and 4, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.
    Plasmid Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 88/100, based on 374 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare ecori cut pgex 4t 1
    VP1 protein expression in E. coli using various expression vectors . The VP1 protein expression in E. coli BL21(DE3) using the expression vectors, pET28a (A) and <t>pGEX-4T-1</t> (B) was analyzed by SDS-PAGE and Western-blotting. Anti-His tag and anti-GST tag monoclonal antibody were respectively used for the recognition of differently tagged VP1 proteins expressed by the different expression vectors used. Lane M, pre-stained protein marker; lane 1 and lane 3, before IPTG induction; lane 2 and 4, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.
    Ecori Cut Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare bacterial expression vectors pgex 4t 1
    VP1 protein expression in E. coli using various expression vectors . The VP1 protein expression in E. coli BL21(DE3) using the expression vectors, pET28a (A) and <t>pGEX-4T-1</t> (B) was analyzed by SDS-PAGE and Western-blotting. Anti-His tag and anti-GST tag monoclonal antibody were respectively used for the recognition of differently tagged VP1 proteins expressed by the different expression vectors used. Lane M, pre-stained protein marker; lane 1 and lane 3, before IPTG induction; lane 2 and 4, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.
    Bacterial Expression Vectors Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare expression vector pgex 4t ­1
    VP1 protein expression in E. coli using various expression vectors . The VP1 protein expression in E. coli BL21(DE3) using the expression vectors, pET28a (A) and <t>pGEX-4T-1</t> (B) was analyzed by SDS-PAGE and Western-blotting. Anti-His tag and anti-GST tag monoclonal antibody were respectively used for the recognition of differently tagged VP1 proteins expressed by the different expression vectors used. Lane M, pre-stained protein marker; lane 1 and lane 3, before IPTG induction; lane 2 and 4, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.
    Expression Vector Pgex 4t ­1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 4t 1 expression plasmid
    Electrophoretic and SDS-PAGE analysis of recombinant pGB2B3. (A) Electrophoretic identification of digested pGB2B3 recombinant (1.0% agarose gel). Lane M, DL2000 marker; lane 1, pGB2B3/BamHI+EcoRI; lane 2, <t>pGEX-4T-1</t> plasmid. (B) Electrophoresis of the PCR product from pGB2B3 (1.0% agarose gel). Lane M, 50 bp marker; lane 1, PCR product of pGB2B3 plasmid; lane 2, PCR product of pGEX-4T-1 plasmid. (C) SDS-PAGE analysis of GST-tagged BD2/3 fusion protein expression. Lane M, protein MW marker; lane 1, expression products of pGEX-4T-1 induced by IPTG; lane 2, expression products of pGB2B3 induced by IPTG; lane 3, supernatant of lysed culture of pGB2B3 induced by IPTG; lane 4, pellet of lysed culture of pGB2B3 induced by IPTG; lane 5, expression products of pGB2B3 without IPTG induction. BD, β-defensin; IPTG, isopropyl-β-D-thiogalactopyranoside; GST, glutathione S transferase.
    Pgex 4t 1 Expression Plasmid, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 93/100, based on 146 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    GE Healthcare glutathione s transferase
    Electrophoretic and SDS-PAGE analysis of recombinant pGB2B3. (A) Electrophoretic identification of digested pGB2B3 recombinant (1.0% agarose gel). Lane M, DL2000 marker; lane 1, pGB2B3/BamHI+EcoRI; lane 2, <t>pGEX-4T-1</t> plasmid. (B) Electrophoresis of the PCR product from pGB2B3 (1.0% agarose gel). Lane M, 50 bp marker; lane 1, PCR product of pGB2B3 plasmid; lane 2, PCR product of pGEX-4T-1 plasmid. (C) SDS-PAGE analysis of GST-tagged BD2/3 fusion protein expression. Lane M, protein MW marker; lane 1, expression products of pGEX-4T-1 induced by IPTG; lane 2, expression products of pGB2B3 induced by IPTG; lane 3, supernatant of lysed culture of pGB2B3 induced by IPTG; lane 4, pellet of lysed culture of pGB2B3 induced by IPTG; lane 5, expression products of pGB2B3 without IPTG induction. BD, β-defensin; IPTG, isopropyl-β-D-thiogalactopyranoside; GST, glutathione S transferase.
    Glutathione S Transferase, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 92/100, based on 857 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare smai digested vector pgex 4t 1
    Electrophoretic and SDS-PAGE analysis of recombinant pGB2B3. (A) Electrophoretic identification of digested pGB2B3 recombinant (1.0% agarose gel). Lane M, DL2000 marker; lane 1, pGB2B3/BamHI+EcoRI; lane 2, <t>pGEX-4T-1</t> plasmid. (B) Electrophoresis of the PCR product from pGB2B3 (1.0% agarose gel). Lane M, 50 bp marker; lane 1, PCR product of pGB2B3 plasmid; lane 2, PCR product of pGEX-4T-1 plasmid. (C) SDS-PAGE analysis of GST-tagged BD2/3 fusion protein expression. Lane M, protein MW marker; lane 1, expression products of pGEX-4T-1 induced by IPTG; lane 2, expression products of pGB2B3 induced by IPTG; lane 3, supernatant of lysed culture of pGB2B3 induced by IPTG; lane 4, pellet of lysed culture of pGB2B3 induced by IPTG; lane 5, expression products of pGB2B3 without IPTG induction. BD, β-defensin; IPTG, isopropyl-β-D-thiogalactopyranoside; GST, glutathione S transferase.
    Smai Digested Vector Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare gst vector pgex 4t 1
    Electrophoretic and SDS-PAGE analysis of recombinant pGB2B3. (A) Electrophoretic identification of digested pGB2B3 recombinant (1.0% agarose gel). Lane M, DL2000 marker; lane 1, pGB2B3/BamHI+EcoRI; lane 2, <t>pGEX-4T-1</t> plasmid. (B) Electrophoresis of the PCR product from pGB2B3 (1.0% agarose gel). Lane M, 50 bp marker; lane 1, PCR product of pGB2B3 plasmid; lane 2, PCR product of pGEX-4T-1 plasmid. (C) SDS-PAGE analysis of GST-tagged BD2/3 fusion protein expression. Lane M, protein MW marker; lane 1, expression products of pGEX-4T-1 induced by IPTG; lane 2, expression products of pGB2B3 induced by IPTG; lane 3, supernatant of lysed culture of pGB2B3 induced by IPTG; lane 4, pellet of lysed culture of pGB2B3 induced by IPTG; lane 5, expression products of pGB2B3 without IPTG induction. BD, β-defensin; IPTG, isopropyl-β-D-thiogalactopyranoside; GST, glutathione S transferase.
    Gst Vector Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare prokaryotic expression vector pgex 4t 1
    (A) Expression and purification of recombinant GST-fused nsp5 analyzed by SDS-PAGE. M, standard protein marker; lane 1, uninduced E.coli transformed with <t>PGEX-4T-1/nsp5;</t> lane 2, induced E.coli transformed with PGEX-4T-1/nsp5; lane 3, uninduced E.coli transformed with PGEX-4T-1; lane 4, induced E. coli transformed with PGEX-4T-1; lane 5, purified GST-nsp5 protein; lane 6, purified GST. (B) Western blotting analysis of purified GST-nsp5. Positive serum to IBV SC021202 were used as the primary antibody. M, standard protein marker; lane 1, purified GST-nsp5; lane 2, GST.
    Prokaryotic Expression Vector Pgex 4t 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 4t 1 gst expression vector
    (A) Expression and purification of recombinant GST-fused nsp5 analyzed by SDS-PAGE. M, standard protein marker; lane 1, uninduced E.coli transformed with <t>PGEX-4T-1/nsp5;</t> lane 2, induced E.coli transformed with PGEX-4T-1/nsp5; lane 3, uninduced E.coli transformed with PGEX-4T-1; lane 4, induced E. coli transformed with PGEX-4T-1; lane 5, purified GST-nsp5 protein; lane 6, purified GST. (B) Western blotting analysis of purified GST-nsp5. Positive serum to IBV SC021202 were used as the primary antibody. M, standard protein marker; lane 1, purified GST-nsp5; lane 2, GST.
    Pgex 4t 1 Gst Expression Vector, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 91/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Cloning strategies for cloning the paracoccin ORF for expression. The left panel shows the standard strategy, with PCR amplification of the largest exon, restriction endonuclease digestion, and cloning into the expression vector pGEX-4T-1. The positions of the exons are displayed on the map of the gene. Genomic DNA template was extracted from P. brasiliensis strain Pb18. Agarose gel electrophoresis (mid-left) shows the corresponding band amplified by PCR. The exon 4 amplicon was cloned by Bam HI and Eco RI digestion. The right panel shows the strategy for synthesis of the predicted paracoccin sequence fused with the 5′-UTR elements for transcription in the vector pUC57. Green arrow, T7 promoter; black box, lacO (lac operator); and red box, the phage T7 trailer sequence for ribosome binding.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Recombinant Paracoccin Reproduces the Biological Properties of the Native Protein and Induces Protective Th1 Immunity against Paracoccidioides brasiliensis Infection

    doi: 10.1371/journal.pntd.0002788

    Figure Lengend Snippet: Cloning strategies for cloning the paracoccin ORF for expression. The left panel shows the standard strategy, with PCR amplification of the largest exon, restriction endonuclease digestion, and cloning into the expression vector pGEX-4T-1. The positions of the exons are displayed on the map of the gene. Genomic DNA template was extracted from P. brasiliensis strain Pb18. Agarose gel electrophoresis (mid-left) shows the corresponding band amplified by PCR. The exon 4 amplicon was cloned by Bam HI and Eco RI digestion. The right panel shows the strategy for synthesis of the predicted paracoccin sequence fused with the 5′-UTR elements for transcription in the vector pUC57. Green arrow, T7 promoter; black box, lacO (lac operator); and red box, the phage T7 trailer sequence for ribosome binding.

    Article Snippet: First, the largest exon was amplified from P. brasiliensis genomic DNA using the oligonucleotide primers FPADG ( 5′-CTGGATCCATGCAAGCACCCGACCAAC-3′ ) and RPADG ( 5′-CGGGAATTCCTACCAACTCGTTATTGATAGAGCGATAA-3′ ), and then cloned into pGEX-4T-1 (GE Healthcare, Upsala, Sweden) flanked by Bam HI and Eco RI restriction sites.

    Techniques: Clone Assay, Expressing, Polymerase Chain Reaction, Amplification, Plasmid Preparation, Agarose Gel Electrophoresis, Sequencing, Binding Assay

    VP1 protein expression in E. coli using various expression vectors . The VP1 protein expression in E. coli BL21(DE3) using the expression vectors, pET28a (A) and pGEX-4T-1 (B) was analyzed by SDS-PAGE and Western-blotting. Anti-His tag and anti-GST tag monoclonal antibody were respectively used for the recognition of differently tagged VP1 proteins expressed by the different expression vectors used. Lane M, pre-stained protein marker; lane 1 and lane 3, before IPTG induction; lane 2 and 4, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.

    Journal: Microbial Cell Factories

    Article Title: High yield expression in a recombinant E. coli of a codon optimized chicken anemia virus capsid protein VP1 useful for vaccine development

    doi: 10.1186/1475-2859-10-56

    Figure Lengend Snippet: VP1 protein expression in E. coli using various expression vectors . The VP1 protein expression in E. coli BL21(DE3) using the expression vectors, pET28a (A) and pGEX-4T-1 (B) was analyzed by SDS-PAGE and Western-blotting. Anti-His tag and anti-GST tag monoclonal antibody were respectively used for the recognition of differently tagged VP1 proteins expressed by the different expression vectors used. Lane M, pre-stained protein marker; lane 1 and lane 3, before IPTG induction; lane 2 and 4, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.

    Article Snippet: Using a specifically designed primer set, forward primer CAV VP1 F: 5'-GCGGAATTCATGGCAAGACGAGCTCGCAGA-3' and reverse primer CAV VP1-R: 5'-GGGCTCGAGTCAGGGCTGCGTCCCCCAGTA-3', respectively containing the EcoR I and Xho I sites, the full-length VP1 gene was amplified using the plasmid pET28a-VP1 as template DNA, and then cloned into the plasmid pGEX-4T-1 (GE healthcare, Piscataway, NJ), which was then designated to be pGEX-4T-1-VP1 (Figure , panel b).

    Techniques: Expressing, SDS Page, Western Blot, Staining, Marker

    Expression of recombinant GST-VP1 protein in three different E. coli strains . The GST-VP1 protein expression in three E. coli strains, BL21(DE3), BL21-CodonPlus(DE3)-RP and BL21(DE3)-pLysS all containing pGEX-4T-1-VP1 was examined. The GST-VP1 protein was examined and detected using SDS-PAGE (A) and Western-blotting (B). Anti-GST tag monoclonal antibody was used to recognize the VP1 protein. Lane M, pre-stained protein marker; lane 1, 4 and 7, before IPTG induction; lane 2, 5 and 8, after IPTG induction for 1 hrs cultivation; lane 3, 6 and 9, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.

    Journal: Microbial Cell Factories

    Article Title: High yield expression in a recombinant E. coli of a codon optimized chicken anemia virus capsid protein VP1 useful for vaccine development

    doi: 10.1186/1475-2859-10-56

    Figure Lengend Snippet: Expression of recombinant GST-VP1 protein in three different E. coli strains . The GST-VP1 protein expression in three E. coli strains, BL21(DE3), BL21-CodonPlus(DE3)-RP and BL21(DE3)-pLysS all containing pGEX-4T-1-VP1 was examined. The GST-VP1 protein was examined and detected using SDS-PAGE (A) and Western-blotting (B). Anti-GST tag monoclonal antibody was used to recognize the VP1 protein. Lane M, pre-stained protein marker; lane 1, 4 and 7, before IPTG induction; lane 2, 5 and 8, after IPTG induction for 1 hrs cultivation; lane 3, 6 and 9, after IPTG induction for 4 h cultivation. The solid triangle pinpoints the expressed VP1 protein.

    Article Snippet: Using a specifically designed primer set, forward primer CAV VP1 F: 5'-GCGGAATTCATGGCAAGACGAGCTCGCAGA-3' and reverse primer CAV VP1-R: 5'-GGGCTCGAGTCAGGGCTGCGTCCCCCAGTA-3', respectively containing the EcoR I and Xho I sites, the full-length VP1 gene was amplified using the plasmid pET28a-VP1 as template DNA, and then cloned into the plasmid pGEX-4T-1 (GE healthcare, Piscataway, NJ), which was then designated to be pGEX-4T-1-VP1 (Figure , panel b).

    Techniques: Expressing, Recombinant, SDS Page, Western Blot, Staining, Marker

    Growth kinetics and production profiles of GST-VP1 protein in three E. coli strains . (A) Growth profile of BL21(DE3), BL21-CodonPlus(DE3)-RP and BL21(DE3)-pLysS in LB medium post-induction. The production profiles of the three E. coli strains, (B)BL21(DE3)-pLysS, (C)BL21(DE3) and (D)BL21-CodonPlus(DE3)-RP containing pGEX-4T-1-VP1 are shown over time course after IPTG induction.

    Journal: Microbial Cell Factories

    Article Title: High yield expression in a recombinant E. coli of a codon optimized chicken anemia virus capsid protein VP1 useful for vaccine development

    doi: 10.1186/1475-2859-10-56

    Figure Lengend Snippet: Growth kinetics and production profiles of GST-VP1 protein in three E. coli strains . (A) Growth profile of BL21(DE3), BL21-CodonPlus(DE3)-RP and BL21(DE3)-pLysS in LB medium post-induction. The production profiles of the three E. coli strains, (B)BL21(DE3)-pLysS, (C)BL21(DE3) and (D)BL21-CodonPlus(DE3)-RP containing pGEX-4T-1-VP1 are shown over time course after IPTG induction.

    Article Snippet: Using a specifically designed primer set, forward primer CAV VP1 F: 5'-GCGGAATTCATGGCAAGACGAGCTCGCAGA-3' and reverse primer CAV VP1-R: 5'-GGGCTCGAGTCAGGGCTGCGTCCCCCAGTA-3', respectively containing the EcoR I and Xho I sites, the full-length VP1 gene was amplified using the plasmid pET28a-VP1 as template DNA, and then cloned into the plasmid pGEX-4T-1 (GE healthcare, Piscataway, NJ), which was then designated to be pGEX-4T-1-VP1 (Figure , panel b).

    Techniques:

    Growth kinetics and production profiles of GST-opt-VP1 protein in E. coli BL21(DE3)-pLysS and BL21(DE3) . (A) Growth profile of BL21(DE3) and BL21(DE3)-pLysS in LB medium during post-induction. (B) The production profiles of the E. coli strains, (B)BL21(DE3)-pLysS, and BL21(DE3) containing pGEX-4T-1-Opt-VP1 are shown after induction.

    Journal: Microbial Cell Factories

    Article Title: High yield expression in a recombinant E. coli of a codon optimized chicken anemia virus capsid protein VP1 useful for vaccine development

    doi: 10.1186/1475-2859-10-56

    Figure Lengend Snippet: Growth kinetics and production profiles of GST-opt-VP1 protein in E. coli BL21(DE3)-pLysS and BL21(DE3) . (A) Growth profile of BL21(DE3) and BL21(DE3)-pLysS in LB medium during post-induction. (B) The production profiles of the E. coli strains, (B)BL21(DE3)-pLysS, and BL21(DE3) containing pGEX-4T-1-Opt-VP1 are shown after induction.

    Article Snippet: Using a specifically designed primer set, forward primer CAV VP1 F: 5'-GCGGAATTCATGGCAAGACGAGCTCGCAGA-3' and reverse primer CAV VP1-R: 5'-GGGCTCGAGTCAGGGCTGCGTCCCCCAGTA-3', respectively containing the EcoR I and Xho I sites, the full-length VP1 gene was amplified using the plasmid pET28a-VP1 as template DNA, and then cloned into the plasmid pGEX-4T-1 (GE healthcare, Piscataway, NJ), which was then designated to be pGEX-4T-1-VP1 (Figure , panel b).

    Techniques:

    Schematic diagram of the constructs used for CVA VP1 protein expression . (A) Schematic representation of the VP1 protein variants and expression vectors used in this study. The designations of the VP1 protein and its expression vectors are indicated, a, b and c. The first two constructs, a and b, contained the full-length VP1 gene cloned into vectors pET28a and pGEX-4T-1, for expression of VP1 protein with a six-histidine (6 × His) tag and glutathione-s-transferase (GST) tag at its N-terminus, respectively. Construct c contained VP1 gene with codon-optimized at its N-terminus; this was derived from construct b by replacing the rare codons in the area 1-321 bp without altering amino acid sequence. The N-terminal codon-optimized VP1 gene, opt-N, was fused with C-terminal domain of VP1 gene (VP1 C-term) by overlapping PCR, and then cloned into pGEX-4T-1. (B) Sequence comparison between the WT VP1 and Opt VP1 gene. The nucleotide sequences were compared between the original VP1 gene (WT VP1) and the sequence of codon-optimized VP1 gene (Opt VP1) over the N-terminal region. An asterisk ( * ) represents the fact that the aligned nucleotides are identical.

    Journal: Microbial Cell Factories

    Article Title: High yield expression in a recombinant E. coli of a codon optimized chicken anemia virus capsid protein VP1 useful for vaccine development

    doi: 10.1186/1475-2859-10-56

    Figure Lengend Snippet: Schematic diagram of the constructs used for CVA VP1 protein expression . (A) Schematic representation of the VP1 protein variants and expression vectors used in this study. The designations of the VP1 protein and its expression vectors are indicated, a, b and c. The first two constructs, a and b, contained the full-length VP1 gene cloned into vectors pET28a and pGEX-4T-1, for expression of VP1 protein with a six-histidine (6 × His) tag and glutathione-s-transferase (GST) tag at its N-terminus, respectively. Construct c contained VP1 gene with codon-optimized at its N-terminus; this was derived from construct b by replacing the rare codons in the area 1-321 bp without altering amino acid sequence. The N-terminal codon-optimized VP1 gene, opt-N, was fused with C-terminal domain of VP1 gene (VP1 C-term) by overlapping PCR, and then cloned into pGEX-4T-1. (B) Sequence comparison between the WT VP1 and Opt VP1 gene. The nucleotide sequences were compared between the original VP1 gene (WT VP1) and the sequence of codon-optimized VP1 gene (Opt VP1) over the N-terminal region. An asterisk ( * ) represents the fact that the aligned nucleotides are identical.

    Article Snippet: Using a specifically designed primer set, forward primer CAV VP1 F: 5'-GCGGAATTCATGGCAAGACGAGCTCGCAGA-3' and reverse primer CAV VP1-R: 5'-GGGCTCGAGTCAGGGCTGCGTCCCCCAGTA-3', respectively containing the EcoR I and Xho I sites, the full-length VP1 gene was amplified using the plasmid pET28a-VP1 as template DNA, and then cloned into the plasmid pGEX-4T-1 (GE healthcare, Piscataway, NJ), which was then designated to be pGEX-4T-1-VP1 (Figure , panel b).

    Techniques: Construct, Expressing, Clone Assay, Derivative Assay, Sequencing, Polymerase Chain Reaction

    Production of five GST fusion proteins, each containing a non-hydrophobic region from the JEV structural protein-coding region. GST fusion proteins were expressed from pGex-4T-1 vector in E . coli and purified from bacterial lysates by affinity chromatography using glutathione-Sepharose. About 5–10 μg of each purified protein was resolved on a SDS-polyacrylamide gel and stained with Coomassie blue. The five GST fusion proteins we generated are: GST-C ( A ), GST-pr ( B ), GST-M ( C ), GST-E N-term ( D ), and GST-E C-term ( E ). Free GST protein was used as control. Molecular weight markers are indicated in kDa on the left side of each panel, and GST and GST fusion proteins are marked on the right side. The predicted molecular weights are indicated at the bottom of each panel.

    Journal: PLoS ONE

    Article Title: Profiling of Viral Proteins Expressed from the Genomic RNA of Japanese Encephalitis Virus Using a Panel of 15 Region-Specific Polyclonal Rabbit Antisera: Implications for Viral Gene Expression

    doi: 10.1371/journal.pone.0124318

    Figure Lengend Snippet: Production of five GST fusion proteins, each containing a non-hydrophobic region from the JEV structural protein-coding region. GST fusion proteins were expressed from pGex-4T-1 vector in E . coli and purified from bacterial lysates by affinity chromatography using glutathione-Sepharose. About 5–10 μg of each purified protein was resolved on a SDS-polyacrylamide gel and stained with Coomassie blue. The five GST fusion proteins we generated are: GST-C ( A ), GST-pr ( B ), GST-M ( C ), GST-E N-term ( D ), and GST-E C-term ( E ). Free GST protein was used as control. Molecular weight markers are indicated in kDa on the left side of each panel, and GST and GST fusion proteins are marked on the right side. The predicted molecular weights are indicated at the bottom of each panel.

    Article Snippet: Next, the resulting PCR amplicons were individually ligated in-frame to the 3'-end of GST coding sequence in the pGex-4T-1 vector (GE Healthcare, Piscataway, NJ) using either the Eco RI and Xho I sites (for the first 14 PCR amplicons except NS5N-term and NS5C-term ) or the Bam HI and Xho I sites (for NS5N-term and NS5C-term ).

    Techniques: Plasmid Preparation, Purification, Affinity Chromatography, Staining, Generated, Molecular Weight

    Electrophoretic and SDS-PAGE analysis of recombinant pGB2B3. (A) Electrophoretic identification of digested pGB2B3 recombinant (1.0% agarose gel). Lane M, DL2000 marker; lane 1, pGB2B3/BamHI+EcoRI; lane 2, pGEX-4T-1 plasmid. (B) Electrophoresis of the PCR product from pGB2B3 (1.0% agarose gel). Lane M, 50 bp marker; lane 1, PCR product of pGB2B3 plasmid; lane 2, PCR product of pGEX-4T-1 plasmid. (C) SDS-PAGE analysis of GST-tagged BD2/3 fusion protein expression. Lane M, protein MW marker; lane 1, expression products of pGEX-4T-1 induced by IPTG; lane 2, expression products of pGB2B3 induced by IPTG; lane 3, supernatant of lysed culture of pGB2B3 induced by IPTG; lane 4, pellet of lysed culture of pGB2B3 induced by IPTG; lane 5, expression products of pGB2B3 without IPTG induction. BD, β-defensin; IPTG, isopropyl-β-D-thiogalactopyranoside; GST, glutathione S transferase.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Improved expression of recombinant fusion defensin gene plasmids packed with chitosan-derived nanoparticles and effect on antibacteria and mouse immunity

    doi: 10.3892/etm.2018.6716

    Figure Lengend Snippet: Electrophoretic and SDS-PAGE analysis of recombinant pGB2B3. (A) Electrophoretic identification of digested pGB2B3 recombinant (1.0% agarose gel). Lane M, DL2000 marker; lane 1, pGB2B3/BamHI+EcoRI; lane 2, pGEX-4T-1 plasmid. (B) Electrophoresis of the PCR product from pGB2B3 (1.0% agarose gel). Lane M, 50 bp marker; lane 1, PCR product of pGB2B3 plasmid; lane 2, PCR product of pGEX-4T-1 plasmid. (C) SDS-PAGE analysis of GST-tagged BD2/3 fusion protein expression. Lane M, protein MW marker; lane 1, expression products of pGEX-4T-1 induced by IPTG; lane 2, expression products of pGB2B3 induced by IPTG; lane 3, supernatant of lysed culture of pGB2B3 induced by IPTG; lane 4, pellet of lysed culture of pGB2B3 induced by IPTG; lane 5, expression products of pGB2B3 without IPTG induction. BD, β-defensin; IPTG, isopropyl-β-D-thiogalactopyranoside; GST, glutathione S transferase.

    Article Snippet: The fusion gene BD2/3 was digested with Bam HI and Eco RI, and then cloned into the pGEX-4T-1 expression plasmid (Pharmacia Biotech; GE Healthcare, Chicago, IL, USA), a prokaryotic expression vector containing the Tac promoter and a glutathione S transferase (GST) tag sequence for fusion protein expression.

    Techniques: SDS Page, Recombinant, Agarose Gel Electrophoresis, Marker, Plasmid Preparation, Electrophoresis, Polymerase Chain Reaction, Expressing

    SDS-PAGE analysis of the expression of pGEX-4T-1/HP-NAP in E. coli . Lane 1: Protein marker; Lane 2: pGEX-4T-1 before induction; Lane 3: pGEX-4T-1 after induction; Lane 4: pGEX-4T-1/HP-NAP before induction; Lane 5: pGEX-4T-1/HP-NAP after induction; Lane

    Journal:

    Article Title: Detection and evaluation of antibodies against neutrophil-activating protein of Helicobacter pylori in patients with gastric cancer

    doi: 10.3748/wjg.15.2381

    Figure Lengend Snippet: SDS-PAGE analysis of the expression of pGEX-4T-1/HP-NAP in E. coli . Lane 1: Protein marker; Lane 2: pGEX-4T-1 before induction; Lane 3: pGEX-4T-1 after induction; Lane 4: pGEX-4T-1/HP-NAP before induction; Lane 5: pGEX-4T-1/HP-NAP after induction; Lane

    Article Snippet: The PCR product was cloned into the expression vector pGEX-4T-1 (Amersham Biosciences).

    Techniques: SDS Page, Expressing, Marker

    Expression of the NTD 231–501 of the PEDV S1 protein in E. coli . (A) The gene region encoding NTD 231–501 was cloned into the pGEX 4T-1 vector containing a GST tag. (B) Recombinant NTD 231–501 of the PEDV S1 protein was expressed by isopropyl β-D-1-thiogalactopyranoside induction and confirmed by Western blot analysis using an anti-GST Ab. (C) Recombinant NTD 231–501 of the PEDV S1 protein was purified by glutathione Sepharose chromatography.

    Journal: Immune Network

    Article Title: N-terminal Domain of the Spike Protein of Porcine Epidemic Diarrhea Virus as a New Candidate Molecule for a Mucosal Vaccine

    doi: 10.4110/in.2018.18.e21

    Figure Lengend Snippet: Expression of the NTD 231–501 of the PEDV S1 protein in E. coli . (A) The gene region encoding NTD 231–501 was cloned into the pGEX 4T-1 vector containing a GST tag. (B) Recombinant NTD 231–501 of the PEDV S1 protein was expressed by isopropyl β-D-1-thiogalactopyranoside induction and confirmed by Western blot analysis using an anti-GST Ab. (C) Recombinant NTD 231–501 of the PEDV S1 protein was purified by glutathione Sepharose chromatography.

    Article Snippet: The amplified NTD231–501 gene was cloned into the pGEX 4T-1 expression vector (GE Healthcare Life Sciences, Buckinghamshire, UK) or the pET SUMO vector (Thermo Fisher Scientific, Waltham, VA, USA).

    Techniques: Expressing, Clone Assay, Plasmid Preparation, Recombinant, Western Blot, Purification, Chromatography

    (A) Expression and purification of recombinant GST-fused nsp5 analyzed by SDS-PAGE. M, standard protein marker; lane 1, uninduced E.coli transformed with PGEX-4T-1/nsp5; lane 2, induced E.coli transformed with PGEX-4T-1/nsp5; lane 3, uninduced E.coli transformed with PGEX-4T-1; lane 4, induced E. coli transformed with PGEX-4T-1; lane 5, purified GST-nsp5 protein; lane 6, purified GST. (B) Western blotting analysis of purified GST-nsp5. Positive serum to IBV SC021202 were used as the primary antibody. M, standard protein marker; lane 1, purified GST-nsp5; lane 2, GST.

    Journal: Journal of Virological Methods

    Article Title: Development and application of nsp5-ELISA for the detection of antibody to infectious bronchitis virus

    doi: 10.1016/j.jviromet.2017.01.026

    Figure Lengend Snippet: (A) Expression and purification of recombinant GST-fused nsp5 analyzed by SDS-PAGE. M, standard protein marker; lane 1, uninduced E.coli transformed with PGEX-4T-1/nsp5; lane 2, induced E.coli transformed with PGEX-4T-1/nsp5; lane 3, uninduced E.coli transformed with PGEX-4T-1; lane 4, induced E. coli transformed with PGEX-4T-1; lane 5, purified GST-nsp5 protein; lane 6, purified GST. (B) Western blotting analysis of purified GST-nsp5. Positive serum to IBV SC021202 were used as the primary antibody. M, standard protein marker; lane 1, purified GST-nsp5; lane 2, GST.

    Article Snippet: The purified PCR product was digested with EcoRI and SalI and cloned into the prokaryotic expression vector pGEX-4T-1 (Amersham, USA).

    Techniques: Expressing, Purification, Recombinant, SDS Page, Marker, Transformation Assay, Western Blot