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  • 99
    GE Healthcare pgex 6p
    Pgex 6p, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 99/100, based on 509 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 99 stars, based on 509 article reviews
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    pgex 6p - by Bioz Stars, 2020-02
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    99
    GE Healthcare pgex 6p 3
    Establishment of monkey podoplanin-neutralizing antibodies (A) Purification of a monkey podoplanin (mkyPDPN) immunogen to establish a hybridoma secreting anti-PDPN monoclonal antibodies (mAbs). A mkyPDPN cDNA region encoding amino acids 76–89 (226–267 bp) was tandemly connected 21 times. The cDNA fragment was inserted into a <t>pGEX-6P-3</t> vector, and a glutathione S-transferase (GST)-tagged mkyPDPN peptide (76–89 aa) produced by BL21 (DE3) E. coli was purified using glutathione sepharose. BALB/c mice were injected with the GST-tagged peptide, after which their splenocytes were fused with mouse myeloma P3U1 cells using polyethylene glycol. Hybridoma screening and antibody purification from ascites were performed. (B) CHO cells transfected with an empty vector (mock), wild-type monkey podoplanin (mkyPDPN-WT), wild-type human podoplanin (hPDPN-WT) were treated with PBS (closed areas) or antibodies (open areas), including anti-PDPN mAb D2-40, 1F6, 2F7, or 3F4 to measure PDPN expression levels. (C) GST-tagged recombinant mkyPDPN protein (WT) and its point mutants were expressed in E. coli . Cell lysates were electrophoresed and immunoblotted with antibodies to PDPN (1F6, 2F7, 3F4) or GST. The PLAG4 domain is indicated by red letters. (D) CHO/mock, CHO/mkyPDPN-WT, or CHO/hPDPN-WT cells were incubated with 100 μg/mL of control IgG1 or anti-PDPN antibodies 1F6, 2F7, or 3F4, followed by incubation with 1 μg/mL of hCLEC-2-(His) 10 (open areas: control IgG-treated samples; green areas: anti-PDPN mAb-treated samples). After washing, cells were further incubated with Alexa Flour 488-conjugated anti-penta-His second antibody. CLEC-2 binding was measured by flow cytometry. Gray areas indicate the fluorescence intensity of samples not treated with CLEC-2. (E) CHO/mkyPDPN-WT cells were incubated with 10 μg/mL of control IgG1, 1F6, 2F7, or 3F4 mAbs followed by incubation with mouse platelet-rich plasma (PRP). The aggregation rate was estimated using an aggregometer. (F) PLAG3 domain-deleted mkyPDPN mutant cells were incubated with 10 μg/mL of control IgG1, 1F6, 2F7, or 3F4 mAbs, followed by incubation with mouse PRP. The aggregation rate was estimated using an aggregometer.
    Pgex 6p 3, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 99/100, based on 624 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p 1
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 99/100, based on 4029 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p 2
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Pgex 6p 2, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 99/100, based on 762 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    GE Healthcare pgex 6p series
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Pgex 6p Series, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 77/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p system
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Pgex 6p System, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 77/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    GE Healthcare bamhi noti pgex 6p 1
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Bamhi Noti Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 78/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p 1 bamhi xhoi
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Pgex 6p 1 Bamhi Xhoi, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 77/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p expression system
    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid <t>pGEX-lepB</t> (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in <t>pGEX-6P-1.</t> Ptac is the tac promoter.
    Pgex 6p Expression System, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 78/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    GE Healthcare pgex 6p 3 plasmid
    Expression and purification of mature CpdB. The recombinant protein was expressed in BL21 cells transformed with <t>pGEX-6P-3-cpdB</t> and induced by IPTG. The GST-CpdB fusion protein was purified by affinity adsorption to GSH-Sepharose. Mature CpdB separated from the GST tag was recovered from the gel after in-column digestion with PreScission. (A) Denaturing gel electrophoresis analysis. L–and L+, bacterial lysates before and after IPTG induction. S and P, supernatant and precipitate of the L+ lysate. M, molecular weight markers (phosphorylase B, 97400; bovine serum albumin, 66200; ovoalbumin, 45000). E, fraction excluded from the GSH-Sepharose column during application of the PreScission protease. 1–4, fractions collected after intracolumn proteolysis. X and Y, bands corresponding to the GST-CpdB fusion and to GPLGS-CpdB obtained by proteolysis. (B) Phosphohydrolytic activities on the indicated substrates, measured in the fractions collected from the GSH-Sepharose column.
    Pgex 6p 3 Plasmid, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 97/100, based on 67 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    GE Healthcare pgex 6p 1 plasmid
    Interaction between LHBsAg and CHC or AP-2. (A) (Top) Schematic representation of the amino acid (a.a.) residues within the LHBsAg subdomains, designated pre-S1, pre-S2, and S; (bottom) Western blot results when LHBsAg cDNA fragments coding for amino acids 1 to 111, 111 to 274, or 274 to 389 were cloned into <t>pGEX-6p-1</t> for expression in E. coli and purification as GST fusion proteins and antibodies against GST were used to detect the expression of the GST fusion proteins. Arrowheads, leaky expression of proteins. (B) GST pulldown assay. GST-LHBsAg fusion proteins or GST bound to glutathione-Sepharose 4B beads were incubated with lysates of HuS-E/2 cells, and then, after GST pulldown, Western blot analysis was performed using antibodies against CHC, AP-1, AP-2, or GST. The positions of molecular mass markers are shown on the left. (C and D) Coimmunoprecipitation and Western blot analysis. HuS-E/2 cells were transfected with plasmid pcDNA3.0-HA-LHBsAg, pcDNA3.0-HA-MHBsAg, or pcDNA3.0-HA-SHBsAg coding, respectively, for HA-tagged LHBsAg, MHBsAg, or SHBsAg, and then, at 2 days posttransfection, the cells were harvested and subjected to immunoprecipitation (IP) with antibodies specific for HA (C) or CHC (D), followed by Western blot analysis with antibodies against HA, AP-2, or CHC, as indicated. NT, nontransfected cells. The molecular mass markers are indicated on the left. Asterisks, proteins coimmunoprecipitated with CHC; arrowheads, nonspecific bands.
    Pgex 6p 1 Plasmid, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 99/100, based on 616 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p 2 plasmid
    Interaction between LHBsAg and CHC or AP-2. (A) (Top) Schematic representation of the amino acid (a.a.) residues within the LHBsAg subdomains, designated pre-S1, pre-S2, and S; (bottom) Western blot results when LHBsAg cDNA fragments coding for amino acids 1 to 111, 111 to 274, or 274 to 389 were cloned into <t>pGEX-6p-1</t> for expression in E. coli and purification as GST fusion proteins and antibodies against GST were used to detect the expression of the GST fusion proteins. Arrowheads, leaky expression of proteins. (B) GST pulldown assay. GST-LHBsAg fusion proteins or GST bound to glutathione-Sepharose 4B beads were incubated with lysates of HuS-E/2 cells, and then, after GST pulldown, Western blot analysis was performed using antibodies against CHC, AP-1, AP-2, or GST. The positions of molecular mass markers are shown on the left. (C and D) Coimmunoprecipitation and Western blot analysis. HuS-E/2 cells were transfected with plasmid pcDNA3.0-HA-LHBsAg, pcDNA3.0-HA-MHBsAg, or pcDNA3.0-HA-SHBsAg coding, respectively, for HA-tagged LHBsAg, MHBsAg, or SHBsAg, and then, at 2 days posttransfection, the cells were harvested and subjected to immunoprecipitation (IP) with antibodies specific for HA (C) or CHC (D), followed by Western blot analysis with antibodies against HA, AP-2, or CHC, as indicated. NT, nontransfected cells. The molecular mass markers are indicated on the left. Asterisks, proteins coimmunoprecipitated with CHC; arrowheads, nonspecific bands.
    Pgex 6p 2 Plasmid, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 96/100, based on 119 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    GE Healthcare bam hi xho i digested pgex 6p 1
    Interaction between LHBsAg and CHC or AP-2. (A) (Top) Schematic representation of the amino acid (a.a.) residues within the LHBsAg subdomains, designated pre-S1, pre-S2, and S; (bottom) Western blot results when LHBsAg cDNA fragments coding for amino acids 1 to 111, 111 to 274, or 274 to 389 were cloned into <t>pGEX-6p-1</t> for expression in E. coli and purification as GST fusion proteins and antibodies against GST were used to detect the expression of the GST fusion proteins. Arrowheads, leaky expression of proteins. (B) GST pulldown assay. GST-LHBsAg fusion proteins or GST bound to glutathione-Sepharose 4B beads were incubated with lysates of HuS-E/2 cells, and then, after GST pulldown, Western blot analysis was performed using antibodies against CHC, AP-1, AP-2, or GST. The positions of molecular mass markers are shown on the left. (C and D) Coimmunoprecipitation and Western blot analysis. HuS-E/2 cells were transfected with plasmid pcDNA3.0-HA-LHBsAg, pcDNA3.0-HA-MHBsAg, or pcDNA3.0-HA-SHBsAg coding, respectively, for HA-tagged LHBsAg, MHBsAg, or SHBsAg, and then, at 2 days posttransfection, the cells were harvested and subjected to immunoprecipitation (IP) with antibodies specific for HA (C) or CHC (D), followed by Western blot analysis with antibodies against HA, AP-2, or CHC, as indicated. NT, nontransfected cells. The molecular mass markers are indicated on the left. Asterisks, proteins coimmunoprecipitated with CHC; arrowheads, nonspecific bands.
    Bam Hi Xho I Digested Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 78/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    GE Healthcare expression vectors pgex 6p 2
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Expression Vectors Pgex 6p 2, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 79/100, based on 154 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    83
    GE Healthcare ecori digested pgex 6p 1 plasmid
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Ecori Digested Pgex 6p 1 Plasmid, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 83/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare ecori sali digested pgex 6p 1
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Ecori Sali Digested Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 79/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare pgex 6p 1 plasmid u78872 1
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Pgex 6p 1 Plasmid U78872 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 80/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare prescission protease pgex 6p
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Prescission Protease Pgex 6p, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 77/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare noti pgex 6p 2 plasmid
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Noti Pgex 6p 2 Plasmid, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 78/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare recombinant pgex 6p 1
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Recombinant Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 97/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    GE Healthcare gst fusion vector pgex 6p 1
    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Gst Fusion Vector Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 86/100, based on 46 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Gateway Compatible Pgex 6p 1 Vector, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 82/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
    Xhoi Digested Pgex 6p 1, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 79/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced <t>pGEX-6P-1-TgPuf1-PUM-HD/BL21.</t> Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).
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    Image Search Results


    Establishment of monkey podoplanin-neutralizing antibodies (A) Purification of a monkey podoplanin (mkyPDPN) immunogen to establish a hybridoma secreting anti-PDPN monoclonal antibodies (mAbs). A mkyPDPN cDNA region encoding amino acids 76–89 (226–267 bp) was tandemly connected 21 times. The cDNA fragment was inserted into a pGEX-6P-3 vector, and a glutathione S-transferase (GST)-tagged mkyPDPN peptide (76–89 aa) produced by BL21 (DE3) E. coli was purified using glutathione sepharose. BALB/c mice were injected with the GST-tagged peptide, after which their splenocytes were fused with mouse myeloma P3U1 cells using polyethylene glycol. Hybridoma screening and antibody purification from ascites were performed. (B) CHO cells transfected with an empty vector (mock), wild-type monkey podoplanin (mkyPDPN-WT), wild-type human podoplanin (hPDPN-WT) were treated with PBS (closed areas) or antibodies (open areas), including anti-PDPN mAb D2-40, 1F6, 2F7, or 3F4 to measure PDPN expression levels. (C) GST-tagged recombinant mkyPDPN protein (WT) and its point mutants were expressed in E. coli . Cell lysates were electrophoresed and immunoblotted with antibodies to PDPN (1F6, 2F7, 3F4) or GST. The PLAG4 domain is indicated by red letters. (D) CHO/mock, CHO/mkyPDPN-WT, or CHO/hPDPN-WT cells were incubated with 100 μg/mL of control IgG1 or anti-PDPN antibodies 1F6, 2F7, or 3F4, followed by incubation with 1 μg/mL of hCLEC-2-(His) 10 (open areas: control IgG-treated samples; green areas: anti-PDPN mAb-treated samples). After washing, cells were further incubated with Alexa Flour 488-conjugated anti-penta-His second antibody. CLEC-2 binding was measured by flow cytometry. Gray areas indicate the fluorescence intensity of samples not treated with CLEC-2. (E) CHO/mkyPDPN-WT cells were incubated with 10 μg/mL of control IgG1, 1F6, 2F7, or 3F4 mAbs followed by incubation with mouse platelet-rich plasma (PRP). The aggregation rate was estimated using an aggregometer. (F) PLAG3 domain-deleted mkyPDPN mutant cells were incubated with 10 μg/mL of control IgG1, 1F6, 2F7, or 3F4 mAbs, followed by incubation with mouse PRP. The aggregation rate was estimated using an aggregometer.

    Journal: Oncotarget

    Article Title: A safety study of newly generated anti-podoplanin-neutralizing antibody in cynomolgus monkey (Macaca fascicularis)

    doi: 10.18632/oncotarget.26055

    Figure Lengend Snippet: Establishment of monkey podoplanin-neutralizing antibodies (A) Purification of a monkey podoplanin (mkyPDPN) immunogen to establish a hybridoma secreting anti-PDPN monoclonal antibodies (mAbs). A mkyPDPN cDNA region encoding amino acids 76–89 (226–267 bp) was tandemly connected 21 times. The cDNA fragment was inserted into a pGEX-6P-3 vector, and a glutathione S-transferase (GST)-tagged mkyPDPN peptide (76–89 aa) produced by BL21 (DE3) E. coli was purified using glutathione sepharose. BALB/c mice were injected with the GST-tagged peptide, after which their splenocytes were fused with mouse myeloma P3U1 cells using polyethylene glycol. Hybridoma screening and antibody purification from ascites were performed. (B) CHO cells transfected with an empty vector (mock), wild-type monkey podoplanin (mkyPDPN-WT), wild-type human podoplanin (hPDPN-WT) were treated with PBS (closed areas) or antibodies (open areas), including anti-PDPN mAb D2-40, 1F6, 2F7, or 3F4 to measure PDPN expression levels. (C) GST-tagged recombinant mkyPDPN protein (WT) and its point mutants were expressed in E. coli . Cell lysates were electrophoresed and immunoblotted with antibodies to PDPN (1F6, 2F7, 3F4) or GST. The PLAG4 domain is indicated by red letters. (D) CHO/mock, CHO/mkyPDPN-WT, or CHO/hPDPN-WT cells were incubated with 100 μg/mL of control IgG1 or anti-PDPN antibodies 1F6, 2F7, or 3F4, followed by incubation with 1 μg/mL of hCLEC-2-(His) 10 (open areas: control IgG-treated samples; green areas: anti-PDPN mAb-treated samples). After washing, cells were further incubated with Alexa Flour 488-conjugated anti-penta-His second antibody. CLEC-2 binding was measured by flow cytometry. Gray areas indicate the fluorescence intensity of samples not treated with CLEC-2. (E) CHO/mkyPDPN-WT cells were incubated with 10 μg/mL of control IgG1, 1F6, 2F7, or 3F4 mAbs followed by incubation with mouse platelet-rich plasma (PRP). The aggregation rate was estimated using an aggregometer. (F) PLAG3 domain-deleted mkyPDPN mutant cells were incubated with 10 μg/mL of control IgG1, 1F6, 2F7, or 3F4 mAbs, followed by incubation with mouse PRP. The aggregation rate was estimated using an aggregometer.

    Article Snippet: The PCR products were subcloned by Topo TA cloning (Invitrogen, Carlsbad, CA, USA) and, after sequence confirmation, cloned into pcDNA3 (Life Technologies, Carlsbad, CA, USA) or pGEX-6P-3 (GE Healthcare, Buckinghamshire, UK) plasmids using EcoRI restriction sites, resulting in pcDNA3 monkey podoplanin or pGEX-6P-3 monkey podoplanin, respectively.

    Techniques: Purification, Plasmid Preparation, Produced, Mouse Assay, Injection, Antibody Purification, Transfection, Expressing, Recombinant, Incubation, Binding Assay, Flow Cytometry, Cytometry, Fluorescence, Mutagenesis

    Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid pGEX-lepB (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in pGEX-6P-1. Ptac is the tac promoter.

    Journal: Journal of Bacteriology

    Article Title: A Second Lysine-Specific Serine Protease from Lysobacter sp. Strain IB-9374

    doi: 10.1128/JB.186.15.5093-5100.2004

    Figure Lengend Snippet: Physical maps of the lepB and lepA genes encoding the lysyl endopeptidases from Lysobacter sp. strain IB-9374 (A) and the expression plasmid pGEX-lepB (B). (A) pLBP5 and pLBS8 are the insert DNAs subcloned into pUC118. The arrows indicate the sizes, directions, and locations of the lepB and lepA genes. The lepB gene is depicted below. The restriction site designations are as follows: B, BamHI; E, EcoRI; P, PstI; Sm, SmaI; Sp, SphI. (B) The partial lepB gene containing the propeptide and mature enzyme regions was amplified by PCR, and the DNA fragment was inserted into BamHI and EcoRI sites downstream of the GST gene in pGEX-6P-1. Ptac is the tac promoter.

    Article Snippet: The insert DNA was digested with BamHI and EcoRI and then ligated into an expression vector of GST fusion protein, pGEX-6P-1 (Amersham Pharmacia Biotech AB), and digested with the same restriction enzyme, giving pGEX-lepB (Fig. ).

    Techniques: Expressing, Plasmid Preparation, Amplification, Polymerase Chain Reaction

    Expression and purification of mature CpdB. The recombinant protein was expressed in BL21 cells transformed with pGEX-6P-3-cpdB and induced by IPTG. The GST-CpdB fusion protein was purified by affinity adsorption to GSH-Sepharose. Mature CpdB separated from the GST tag was recovered from the gel after in-column digestion with PreScission. (A) Denaturing gel electrophoresis analysis. L–and L+, bacterial lysates before and after IPTG induction. S and P, supernatant and precipitate of the L+ lysate. M, molecular weight markers (phosphorylase B, 97400; bovine serum albumin, 66200; ovoalbumin, 45000). E, fraction excluded from the GSH-Sepharose column during application of the PreScission protease. 1–4, fractions collected after intracolumn proteolysis. X and Y, bands corresponding to the GST-CpdB fusion and to GPLGS-CpdB obtained by proteolysis. (B) Phosphohydrolytic activities on the indicated substrates, measured in the fractions collected from the GSH-Sepharose column.

    Journal: PLoS ONE

    Article Title: The Characterization of Escherichia coli CpdB as a Recombinant Protein Reveals that, besides Having the Expected 3´-Nucleotidase and 2´,3´-Cyclic Mononucleotide Phosphodiesterase Activities, It Is Also Active as Cyclic Dinucleotide Phosphodiesterase

    doi: 10.1371/journal.pone.0157308

    Figure Lengend Snippet: Expression and purification of mature CpdB. The recombinant protein was expressed in BL21 cells transformed with pGEX-6P-3-cpdB and induced by IPTG. The GST-CpdB fusion protein was purified by affinity adsorption to GSH-Sepharose. Mature CpdB separated from the GST tag was recovered from the gel after in-column digestion with PreScission. (A) Denaturing gel electrophoresis analysis. L–and L+, bacterial lysates before and after IPTG induction. S and P, supernatant and precipitate of the L+ lysate. M, molecular weight markers (phosphorylase B, 97400; bovine serum albumin, 66200; ovoalbumin, 45000). E, fraction excluded from the GSH-Sepharose column during application of the PreScission protease. 1–4, fractions collected after intracolumn proteolysis. X and Y, bands corresponding to the GST-CpdB fusion and to GPLGS-CpdB obtained by proteolysis. (B) Phosphohydrolytic activities on the indicated substrates, measured in the fractions collected from the GSH-Sepharose column.

    Article Snippet: The expression plasmid pGEX-6P-3-cpdB was constructed by directionally subcloning the BamHI-EcoRI insert obtained from pGEM-T-Easy-cpdB between the corresponding sites of plasmid pGEX-6P-3 (GE Healthcare Life Sciences), and transforming JM109 cells with the ligation mixture.

    Techniques: Expressing, Purification, Recombinant, Transformation Assay, Adsorption, Nucleic Acid Electrophoresis, Molecular Weight

    Interaction between LHBsAg and CHC or AP-2. (A) (Top) Schematic representation of the amino acid (a.a.) residues within the LHBsAg subdomains, designated pre-S1, pre-S2, and S; (bottom) Western blot results when LHBsAg cDNA fragments coding for amino acids 1 to 111, 111 to 274, or 274 to 389 were cloned into pGEX-6p-1 for expression in E. coli and purification as GST fusion proteins and antibodies against GST were used to detect the expression of the GST fusion proteins. Arrowheads, leaky expression of proteins. (B) GST pulldown assay. GST-LHBsAg fusion proteins or GST bound to glutathione-Sepharose 4B beads were incubated with lysates of HuS-E/2 cells, and then, after GST pulldown, Western blot analysis was performed using antibodies against CHC, AP-1, AP-2, or GST. The positions of molecular mass markers are shown on the left. (C and D) Coimmunoprecipitation and Western blot analysis. HuS-E/2 cells were transfected with plasmid pcDNA3.0-HA-LHBsAg, pcDNA3.0-HA-MHBsAg, or pcDNA3.0-HA-SHBsAg coding, respectively, for HA-tagged LHBsAg, MHBsAg, or SHBsAg, and then, at 2 days posttransfection, the cells were harvested and subjected to immunoprecipitation (IP) with antibodies specific for HA (C) or CHC (D), followed by Western blot analysis with antibodies against HA, AP-2, or CHC, as indicated. NT, nontransfected cells. The molecular mass markers are indicated on the left. Asterisks, proteins coimmunoprecipitated with CHC; arrowheads, nonspecific bands.

    Journal: Journal of Virology

    Article Title: Entry of Hepatitis B Virus into Immortalized Human Primary Hepatocytes by Clathrin-Dependent Endocytosis

    doi: 10.1128/JVI.00873-12

    Figure Lengend Snippet: Interaction between LHBsAg and CHC or AP-2. (A) (Top) Schematic representation of the amino acid (a.a.) residues within the LHBsAg subdomains, designated pre-S1, pre-S2, and S; (bottom) Western blot results when LHBsAg cDNA fragments coding for amino acids 1 to 111, 111 to 274, or 274 to 389 were cloned into pGEX-6p-1 for expression in E. coli and purification as GST fusion proteins and antibodies against GST were used to detect the expression of the GST fusion proteins. Arrowheads, leaky expression of proteins. (B) GST pulldown assay. GST-LHBsAg fusion proteins or GST bound to glutathione-Sepharose 4B beads were incubated with lysates of HuS-E/2 cells, and then, after GST pulldown, Western blot analysis was performed using antibodies against CHC, AP-1, AP-2, or GST. The positions of molecular mass markers are shown on the left. (C and D) Coimmunoprecipitation and Western blot analysis. HuS-E/2 cells were transfected with plasmid pcDNA3.0-HA-LHBsAg, pcDNA3.0-HA-MHBsAg, or pcDNA3.0-HA-SHBsAg coding, respectively, for HA-tagged LHBsAg, MHBsAg, or SHBsAg, and then, at 2 days posttransfection, the cells were harvested and subjected to immunoprecipitation (IP) with antibodies specific for HA (C) or CHC (D), followed by Western blot analysis with antibodies against HA, AP-2, or CHC, as indicated. NT, nontransfected cells. The molecular mass markers are indicated on the left. Asterisks, proteins coimmunoprecipitated with CHC; arrowheads, nonspecific bands.

    Article Snippet: To generate plasmids pGEX-6p-1-LHBsAg(1-111), pGEX-6p-1-LHBsAg(111-274), and pGEX-6p-1-LHBsAg(274-389), coding for glutathione S -transferase (GST)–LHBsAg1-111 , GST–LHBsAg111-274 , and GST–LHBsAg274-389 , respectively, a KpnI/EcoRI fragment (394 bp), EcoRI/BamHI fragment (489 bp), or BamHI/ApaI fragment (323 bp) was obtained from pcDNA3.0-HA-LHBsAg and subcloned, respectively, into the SmaI, SalI, or BamHI/SmaI site of plasmid pGEX-6P-1 (GE Healthcare Biosciences) following a blunt-end reaction.

    Techniques: Western Blot, Clone Assay, Expressing, Purification, GST Pulldown Assay, Incubation, Transfection, Plasmid Preparation, Immunoprecipitation

    Molecular characterization of 35S::TKTKK3 transgenic plants and western blot hybridization. (a,b) show the bar diagrams of expression patterns of both genes TKTKK1 and TKTKK2 in 55 35S::TKTKK3 transgenic plants by qRT-PCR, respectively. (c) Southern blot hybridization for detecting copy numbers of T-DNA insertion. DNA samples (line numbers were labelled on the top of the panel) were digested by the restriction enzyme Eco RV and were then transferred into nylon member for hybridization with the HPT probe. The red stars “*” indicated the lines with single copy number of T-DNA insertion. (d,e) Western blot hybridization using crude proteins from E. coli (d) and T2 rice grains (e) , respectively. The “M” in (d,e) indicates the protein marker used for the hybridization. The numbers in (d) indicate the crude proteins extracted from E. coli carrying the plasmid pGEX-6P-1 (1 and 2) and pGEX-6P-1::TKTKK1 (3 and 4). The white arrows indicated the GST expression in the E. coli carrying the plasmid pGEX-6P-1 . The green arrows indicated the GST fusion protein expression in the E. coli carrying pGEX-6P-1::TKTKK1 . The numbers in (e) indicate the different transgenic lines used for crude proteins extraction from rice seeds. Three lines (1, line 9, 2, line 14 and 3, line 21) were from 35S::TKTKK1 . Line 46 (indicated by 4) and Line 5 (indicated by 5) were from 35S::TKTKK2 and 35S::TKTKK3 , respectively. The number 6 indicates WT and 7 indicates negative control. The green arrows indicated the stable expression of TKTKK1 protein in lines 9, 14 and 21 by Western blot hybridization.

    Journal: Scientific Reports

    Article Title: Improving protein content and quality by over-expressing artificially synthetic fusion proteins with high lysine and threonine constituent in rice plants

    doi: 10.1038/srep34427

    Figure Lengend Snippet: Molecular characterization of 35S::TKTKK3 transgenic plants and western blot hybridization. (a,b) show the bar diagrams of expression patterns of both genes TKTKK1 and TKTKK2 in 55 35S::TKTKK3 transgenic plants by qRT-PCR, respectively. (c) Southern blot hybridization for detecting copy numbers of T-DNA insertion. DNA samples (line numbers were labelled on the top of the panel) were digested by the restriction enzyme Eco RV and were then transferred into nylon member for hybridization with the HPT probe. The red stars “*” indicated the lines with single copy number of T-DNA insertion. (d,e) Western blot hybridization using crude proteins from E. coli (d) and T2 rice grains (e) , respectively. The “M” in (d,e) indicates the protein marker used for the hybridization. The numbers in (d) indicate the crude proteins extracted from E. coli carrying the plasmid pGEX-6P-1 (1 and 2) and pGEX-6P-1::TKTKK1 (3 and 4). The white arrows indicated the GST expression in the E. coli carrying the plasmid pGEX-6P-1 . The green arrows indicated the GST fusion protein expression in the E. coli carrying pGEX-6P-1::TKTKK1 . The numbers in (e) indicate the different transgenic lines used for crude proteins extraction from rice seeds. Three lines (1, line 9, 2, line 14 and 3, line 21) were from 35S::TKTKK1 . Line 46 (indicated by 4) and Line 5 (indicated by 5) were from 35S::TKTKK2 and 35S::TKTKK3 , respectively. The number 6 indicates WT and 7 indicates negative control. The green arrows indicated the stable expression of TKTKK1 protein in lines 9, 14 and 21 by Western blot hybridization.

    Article Snippet: GST-tagged TKTKK1 construction and Western blot hybridization The GST -tagged pGEX-6P-1 vector (GE Healthcare Life Sciences) was used for sub-cloning TKTKK1 by fusing with the GST sequence at its 3′-terminal.

    Techniques: Transgenic Assay, Western Blot, Hybridization, Expressing, Quantitative RT-PCR, Southern Blot, Marker, Plasmid Preparation, Negative Control

    SDS-PAGE analysis of purified EstS protein. M: Protein molecular weight marker; 1: Uninduced cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-1; 2: IPTG-induced of cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-1; 3: Uninduced cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-estS; 4: IPTG-induced of cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-estS; 5: Purified EstS. The protein GST-EstS is indicated by arrow

    Journal: BMC Biotechnology

    Article Title: Characterization of a cold-active esterase from Serratia sp. and improvement of thermostability by directed evolution

    doi: 10.1186/s12896-016-0235-3

    Figure Lengend Snippet: SDS-PAGE analysis of purified EstS protein. M: Protein molecular weight marker; 1: Uninduced cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-1; 2: IPTG-induced of cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-1; 3: Uninduced cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-estS; 4: IPTG-induced of cell lysate of E. coli BL21 (DE3) harboring pGEX-6P-estS; 5: Purified EstS. The protein GST-EstS is indicated by arrow

    Article Snippet: The vector pGEX-6P-1 (GE Healthcare, USA) was used for gene cloning and protein expression.

    Techniques: SDS Page, Purification, Molecular Weight, Marker

    Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced pGEX-6P-1-TgPuf1-PUM-HD/BL21. Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).

    Journal: Parasites & Vectors

    Article Title: Characterization of TgPuf1, a member of the Puf family RNA-binding proteins from Toxoplasma gondii

    doi: 10.1186/1756-3305-7-141

    Figure Lengend Snippet: Expression and purification of the rTgPuf1 Puf domain in E. coli . (A) Left panel shows a Coomassie Blue stained gel. Lane 1 and 2 shows the pellet and supernatant of lysates of induced pGEX-6P-1-TgPuf1-PUM-HD/BL21. Right panel is the immunoblot with anti-GST antibodies, which detected rTgPuf1 expression in uninduced (lane 3) and IPTG induced BL21 cells (lane 4). (B) Purification of rTgPuf1. Lane 1, lysate of induced BL21 cells; lane 2, lysate passed through a GST column; lane 3–7, elution with 50 mM Tris–HCl and 10 mM glutathione (pH 8.0).

    Article Snippet: Expression of recombinant TgPuf1 Puf domain in Escherichia coli To express the conserved RNA-binding domain of TgPuf1 in bacteria, PCR was performed with T. gondii cDNA using two primers (CGGGATCC AGAAAAGGCGACTCAAAAG and ATAAGAATGCGGCCGC GTCACTGAAACCTGAGATG) designed to clone at the Bam HI and Not I sites of the expression vector pGEX-6P-1 (GE Healthcare).

    Techniques: Expressing, Purification, Staining