iptg Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Thermo Fisher isopropyl thiogalactoside iptg
    Isopropyl Thiogalactoside Iptg, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 82 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/isopropyl thiogalactoside iptg/product/Thermo Fisher
    Average 99 stars, based on 82 article reviews
    Price from $9.99 to $1999.99
    isopropyl thiogalactoside iptg - by Bioz Stars, 2020-07
    99/100 stars
      Buy from Supplier

    99
    Millipore isopropyl thiogalactoside iptg
    Bacterial expression of recombinant PfPP5. Soluble S100 extract (approximately 30 μg protein) of <t>IPTG-induced</t> (lanes 2, 4) or uninduced (lanes 1, 3) E. coli <t>BL21(DE3)-RIG</t> containing pET-15b-PfPP5 (lanes 1, 2) or pET-15b-ΔTPR-PfPP5 (lanes 3, 4) were analyzed by SDS-PAGE followed by staining of the gel with Coomassie Brilliant Blue R250 [ 12 ]. The respective purified proteins were analyzed in lanes 5, 6. Lane 7 represents about 5 μg of purified His-tagged PfPP5 digested with 50 ng of trypsin for 30 min at 30°. Protein markers (lane M) are indicated by their Mr in thousands.
    Isopropyl Thiogalactoside Iptg, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1222 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/isopropyl thiogalactoside iptg/product/Millipore
    Average 99 stars, based on 1222 article reviews
    Price from $9.99 to $1999.99
    isopropyl thiogalactoside iptg - by Bioz Stars, 2020-07
    99/100 stars
      Buy from Supplier

    88
    Research Products International isopropyl thiogalactoside iptg
    Bacterial expression of recombinant PfPP5. Soluble S100 extract (approximately 30 μg protein) of <t>IPTG-induced</t> (lanes 2, 4) or uninduced (lanes 1, 3) E. coli <t>BL21(DE3)-RIG</t> containing pET-15b-PfPP5 (lanes 1, 2) or pET-15b-ΔTPR-PfPP5 (lanes 3, 4) were analyzed by SDS-PAGE followed by staining of the gel with Coomassie Brilliant Blue R250 [ 12 ]. The respective purified proteins were analyzed in lanes 5, 6. Lane 7 represents about 5 μg of purified His-tagged PfPP5 digested with 50 ng of trypsin for 30 min at 30°. Protein markers (lane M) are indicated by their Mr in thousands.
    Isopropyl Thiogalactoside Iptg, supplied by Research Products International, used in various techniques. Bioz Stars score: 88/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/isopropyl thiogalactoside iptg/product/Research Products International
    Average 88 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    isopropyl thiogalactoside iptg - by Bioz Stars, 2020-07
    88/100 stars
      Buy from Supplier

    92
    Denville Scientific iptg
    Bacterial expression of recombinant PfPP5. Soluble S100 extract (approximately 30 μg protein) of <t>IPTG-induced</t> (lanes 2, 4) or uninduced (lanes 1, 3) E. coli <t>BL21(DE3)-RIG</t> containing pET-15b-PfPP5 (lanes 1, 2) or pET-15b-ΔTPR-PfPP5 (lanes 3, 4) were analyzed by SDS-PAGE followed by staining of the gel with Coomassie Brilliant Blue R250 [ 12 ]. The respective purified proteins were analyzed in lanes 5, 6. Lane 7 represents about 5 μg of purified His-tagged PfPP5 digested with 50 ng of trypsin for 30 min at 30°. Protein markers (lane M) are indicated by their Mr in thousands.
    Iptg, supplied by Denville Scientific, used in various techniques. Bioz Stars score: 92/100, based on 97 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/iptg/product/Denville Scientific
    Average 92 stars, based on 97 article reviews
    Price from $9.99 to $1999.99
    iptg - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    88
    GE Healthcare isopropyl thiogalactoside iptg
    Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of recombinant interleukin-6 (rIL-6) ( A ) and recombinant outer membrane protein V (rOmpV) ( B ). Lane M: protein marker; Lanes 1and 4: recombinant clones without isopropy-β- d <t>-thiogalactoside</t> <t>(IPTG)</t> induction; Lanes 2 and 5: recombinant clones with IPTG induction; Lanes 3 and 6: purified rIL-6 and rOmpV.
    Isopropyl Thiogalactoside Iptg, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 88/100, based on 32 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/isopropyl thiogalactoside iptg/product/GE Healthcare
    Average 88 stars, based on 32 article reviews
    Price from $9.99 to $1999.99
    isopropyl thiogalactoside iptg - by Bioz Stars, 2020-07
    88/100 stars
      Buy from Supplier

    95
    Millipore β isopropyl thiogalactoside
    Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of recombinant interleukin-6 (rIL-6) ( A ) and recombinant outer membrane protein V (rOmpV) ( B ). Lane M: protein marker; Lanes 1and 4: recombinant clones without isopropy-β- d <t>-thiogalactoside</t> <t>(IPTG)</t> induction; Lanes 2 and 5: recombinant clones with IPTG induction; Lanes 3 and 6: purified rIL-6 and rOmpV.
    β Isopropyl Thiogalactoside, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β isopropyl thiogalactoside/product/Millipore
    Average 95 stars, based on 24 article reviews
    Price from $9.99 to $1999.99
    β isopropyl thiogalactoside - by Bioz Stars, 2020-07
    95/100 stars
      Buy from Supplier

    93
    Promega iptg
    Cyclin D1 is not required for p14ARF-induced cell growth (size, [forward scatter; FSC-A]). MCF7 cells were transfected in triplicate with siRNA as described in Figure 4 , and cells were harvested at day 3 and analysed using a BD LSRII cytometer. Flow cytometry data (20,000 events) were acquired using DIVA software (version 6.1.3), analysed with CellQuest Pro (version 6) and composite figures were generated with CANVAS (version X) software. A. To eliminate cell doublets and clumps from the analysis, single cells were gated (R1) from FSC-A versus <t>FCS-H</t> dot plots. A representative plot only is shown from mock-transfected cells. FSC-A histograms of the R1 populations were then used to indicate relative cell size of MCF-7 cells ± <t>IPTG</t> treatment after siRNA knockdown of cyclin D1, p53 and p21. Black line shows cells without IPTG and the red line shows cells treated with IPTG. B. Mean Fluorescent Intensity (MFI) analysis of FSC-A from triplicate biological experiments; data shown are mean MFI ± SD. C. Cell counts were performed using a haemocytometer. Data were presented as mean cell counts (x10 4 ) ± SD performed on three separate occasions.
    Iptg, supplied by Promega, used in various techniques. Bioz Stars score: 93/100, based on 824 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/iptg/product/Promega
    Average 93 stars, based on 824 article reviews
    Price from $9.99 to $1999.99
    iptg - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    iptg  (TaKaRa)
    97
    TaKaRa iptg
    3F3A mAb, but not anti-Gp78C, selectively recognizes dephosphorylated <t>Gp78.</t> (A) Cos7 cells were untransfected (Cos7 NT) or transfected with empty vector (PC3 EV), FLAG-tagged wild type (WT), S538A, phosphomimetic S538D or S538E, D536A mutant, or amino acid 533–541 deletion mutant (Δ3F3A) of Gp78. Cell lysates were immunoprecipitated with FLAG beads and probed with 3F3A, Gp78C, or FLAG mAbs. (B) Bacteria transformed with GST-tagged empty vector (GST-EV) or Gp78 C-terminal (GST-Gp78C) wild type (WT), S538A, S538D, S538E mutants, or amino acid 533–541 deletion mutant (Δ3F3A) were uninduced or induced by <t>IPTG.</t> Total bacterial lysates were dot blotted onto membranes and probed with 3F3A, Gp78C, or GST mAbs. (C) FLAG-tagged wild type of Gp78 (FLAG-Gp78-WT) or phosphorylation dominant-negative mutant (FLAG-Gp78 S538A) from Cos7 cells were immunoprecipitated and treated or not with CIAP and probed with 3F3A mAb, Gp78C, or FLAG mAb (±SEM; n = 3).
    Iptg, supplied by TaKaRa, used in various techniques. Bioz Stars score: 97/100, based on 756 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/iptg/product/TaKaRa
    Average 97 stars, based on 756 article reviews
    Price from $9.99 to $1999.99
    iptg - by Bioz Stars, 2020-07
    97/100 stars
      Buy from Supplier

    99
    Thermo Fisher iptg
    Fig. 2. Electrophoretic mobility and purification of HEβ. ( A ) Western blot developed with anti-E-tag mAb–POD of whole-cell protein extracts from <t>IPTG-induced</t> E.coli UT5600 harboring pHEβ. Before loading onto the 10% polyacrylamide gel, the samples were <t>resuspended</t> in denaturing SDS–PAGE sample buffer and heated for 10 min at the indicated temperatures (25, 42, 50, 65 and 100°C). The faster mobility band of HEβ (f) corresponds to the folded conformation of the hybrid. When unfolded, HEβ migrates as a slower mobility band (u). ( B ) A sample of purified HEβ is shown after Coomassie Blue staining of a 10% SDS–polyacrylamide gel. ( C ) Western blot developed with anti- E-tag mAb–POD of purified HEβ samples treated at 25 (lane 1) or 100°C (lane 2) for 10 min before loading onto a 10% SDS– polyacrylamide gel. ( D ) The possible presence of contaminating OmpF porin in the purified HEβ was evaluated by western blot developed with a polyclonal serum against trimeric OmpF. Excess purified HEβ (10 µg, lanes 1 and 2) and a sample of purified OmpF (0.1 µg, lanes 3 and 4) as a control were loaded onto a 10% SDS–polyacrylamide gel after heating at 25 (lanes 1 and 3) or 100°C (lanes 2 and 4) for 10 min. Only the trimeric OmpF control (lane 3) was detected, ruling out the presence of OmpF in the purified HEβ sample.
    Iptg, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 4961 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/iptg/product/Thermo Fisher
    Average 99 stars, based on 4961 article reviews
    Price from $9.99 to $1999.99
    iptg - by Bioz Stars, 2020-07
    99/100 stars
      Buy from Supplier

    iptg  (Abcam)
    94
    Abcam iptg
    Fig. 2. Electrophoretic mobility and purification of HEβ. ( A ) Western blot developed with anti-E-tag mAb–POD of whole-cell protein extracts from <t>IPTG-induced</t> E.coli UT5600 harboring pHEβ. Before loading onto the 10% polyacrylamide gel, the samples were <t>resuspended</t> in denaturing SDS–PAGE sample buffer and heated for 10 min at the indicated temperatures (25, 42, 50, 65 and 100°C). The faster mobility band of HEβ (f) corresponds to the folded conformation of the hybrid. When unfolded, HEβ migrates as a slower mobility band (u). ( B ) A sample of purified HEβ is shown after Coomassie Blue staining of a 10% SDS–polyacrylamide gel. ( C ) Western blot developed with anti- E-tag mAb–POD of purified HEβ samples treated at 25 (lane 1) or 100°C (lane 2) for 10 min before loading onto a 10% SDS– polyacrylamide gel. ( D ) The possible presence of contaminating OmpF porin in the purified HEβ was evaluated by western blot developed with a polyclonal serum against trimeric OmpF. Excess purified HEβ (10 µg, lanes 1 and 2) and a sample of purified OmpF (0.1 µg, lanes 3 and 4) as a control were loaded onto a 10% SDS–polyacrylamide gel after heating at 25 (lanes 1 and 3) or 100°C (lanes 2 and 4) for 10 min. Only the trimeric OmpF control (lane 3) was detected, ruling out the presence of OmpF in the purified HEβ sample.
    Iptg, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 41 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/iptg/product/Abcam
    Average 94 stars, based on 41 article reviews
    Price from $9.99 to $1999.99
    iptg - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    Image Search Results


    Bacterial expression of recombinant PfPP5. Soluble S100 extract (approximately 30 μg protein) of IPTG-induced (lanes 2, 4) or uninduced (lanes 1, 3) E. coli BL21(DE3)-RIG containing pET-15b-PfPP5 (lanes 1, 2) or pET-15b-ΔTPR-PfPP5 (lanes 3, 4) were analyzed by SDS-PAGE followed by staining of the gel with Coomassie Brilliant Blue R250 [ 12 ]. The respective purified proteins were analyzed in lanes 5, 6. Lane 7 represents about 5 μg of purified His-tagged PfPP5 digested with 50 ng of trypsin for 30 min at 30°. Protein markers (lane M) are indicated by their Mr in thousands.

    Journal: BMC Microbiology

    Article Title: A novel tetratricopeptide repeat (TPR) containing PP5 serine/threonine protein phosphatase in the malaria parasite, Plasmodium falciparum

    doi:

    Figure Lengend Snippet: Bacterial expression of recombinant PfPP5. Soluble S100 extract (approximately 30 μg protein) of IPTG-induced (lanes 2, 4) or uninduced (lanes 1, 3) E. coli BL21(DE3)-RIG containing pET-15b-PfPP5 (lanes 1, 2) or pET-15b-ΔTPR-PfPP5 (lanes 3, 4) were analyzed by SDS-PAGE followed by staining of the gel with Coomassie Brilliant Blue R250 [ 12 ]. The respective purified proteins were analyzed in lanes 5, 6. Lane 7 represents about 5 μg of purified His-tagged PfPP5 digested with 50 ng of trypsin for 30 min at 30°. Protein markers (lane M) are indicated by their Mr in thousands.

    Article Snippet: Introduction of the plasmids into E. coli BL21(DE3) cells containing the compatible RIG plasmid [ ], induction of the proteins with IPTG, and purification through Ni-chelation chromatography were performed using the His-Bind kit reagents and procedure (Novagen).

    Techniques: Expressing, Recombinant, Positron Emission Tomography, SDS Page, Staining, Purification

    Expression dynamics of SGIV VP19. (A) SDS-PAGE analysis of purified recombinant SGIV VP19. Lines M, 1, 2, 3, 4 and 5 showed protein markers, pET-VP19 (uninduced), pET-VP19 (IPTG induced), supernatant, pellet of induced pET-VP19 and the purified pET-VP19, respectively. (B) The specificity of anti-VP19 serum. SGIV or mock infected cells at 48 h p.i. were collected and centrifuged for SDS-PAGE and western blotting. Line M, 1 and 2 showed protein markers, mock and SGIV infected cells at 48 h p.i., respectively. (C) The expression pattern of VP19 during SGIV infection. Actin was chosen as the internal control. (D) VP19 was identified as a late protein. Cells were infected with SGIV at MOI of 0.5 for 6 h and 24 h under the treatment with CHX or AraC, respectively. Then cells were collected for western blotting analysis. The late protein MCP was chosen as a positive control.

    Journal: Virology Journal

    Article Title: Characterization of an envelope gene VP19 from Singapore grouper iridovirus

    doi: 10.1186/1743-422X-10-354

    Figure Lengend Snippet: Expression dynamics of SGIV VP19. (A) SDS-PAGE analysis of purified recombinant SGIV VP19. Lines M, 1, 2, 3, 4 and 5 showed protein markers, pET-VP19 (uninduced), pET-VP19 (IPTG induced), supernatant, pellet of induced pET-VP19 and the purified pET-VP19, respectively. (B) The specificity of anti-VP19 serum. SGIV or mock infected cells at 48 h p.i. were collected and centrifuged for SDS-PAGE and western blotting. Line M, 1 and 2 showed protein markers, mock and SGIV infected cells at 48 h p.i., respectively. (C) The expression pattern of VP19 during SGIV infection. Actin was chosen as the internal control. (D) VP19 was identified as a late protein. Cells were infected with SGIV at MOI of 0.5 for 6 h and 24 h under the treatment with CHX or AraC, respectively. Then cells were collected for western blotting analysis. The late protein MCP was chosen as a positive control.

    Article Snippet: After IPTG induction of E. coli BL21 (DE3) containing pET-VP19t, the recombinant fusion protein rVP19t was purified using the HisBind purification kit (Novagen, Germany) according to the manufacture’s protocol.

    Techniques: Expressing, SDS Page, Purification, Recombinant, Positron Emission Tomography, Infection, Western Blot, Positive Control

    Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of recombinant interleukin-6 (rIL-6) ( A ) and recombinant outer membrane protein V (rOmpV) ( B ). Lane M: protein marker; Lanes 1and 4: recombinant clones without isopropy-β- d -thiogalactoside (IPTG) induction; Lanes 2 and 5: recombinant clones with IPTG induction; Lanes 3 and 6: purified rIL-6 and rOmpV.

    Journal: International Journal of Molecular Sciences

    Article Title: The Immune Adjuvant Effects of Flounder (Paralichthys olivaceus) Interleukin-6 on E. tarda Subunit Vaccine OmpV

    doi: 10.3390/ijms18071445

    Figure Lengend Snippet: Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of recombinant interleukin-6 (rIL-6) ( A ) and recombinant outer membrane protein V (rOmpV) ( B ). Lane M: protein marker; Lanes 1and 4: recombinant clones without isopropy-β- d -thiogalactoside (IPTG) induction; Lanes 2 and 5: recombinant clones with IPTG induction; Lanes 3 and 6: purified rIL-6 and rOmpV.

    Article Snippet: When the OD600 of the cultures reached 0.6, 1 mM isopropyl thiogalactoside (IPTG) was added and grown at 37 °C for an additional 10 h. Then, the cultures were centrifuged, and the His-tagged rIL-6 was purified using His TrapTM HP Ni-Agarose (GE healthcare, Beijing, China).

    Techniques: Polyacrylamide Gel Electrophoresis, SDS Page, Recombinant, Marker, Clone Assay, Purification

    Cyclin D1 is not required for p14ARF-induced cell growth (size, [forward scatter; FSC-A]). MCF7 cells were transfected in triplicate with siRNA as described in Figure 4 , and cells were harvested at day 3 and analysed using a BD LSRII cytometer. Flow cytometry data (20,000 events) were acquired using DIVA software (version 6.1.3), analysed with CellQuest Pro (version 6) and composite figures were generated with CANVAS (version X) software. A. To eliminate cell doublets and clumps from the analysis, single cells were gated (R1) from FSC-A versus FCS-H dot plots. A representative plot only is shown from mock-transfected cells. FSC-A histograms of the R1 populations were then used to indicate relative cell size of MCF-7 cells ± IPTG treatment after siRNA knockdown of cyclin D1, p53 and p21. Black line shows cells without IPTG and the red line shows cells treated with IPTG. B. Mean Fluorescent Intensity (MFI) analysis of FSC-A from triplicate biological experiments; data shown are mean MFI ± SD. C. Cell counts were performed using a haemocytometer. Data were presented as mean cell counts (x10 4 ) ± SD performed on three separate occasions.

    Journal: PLoS ONE

    Article Title: p14ARF Post-Transcriptional Regulation of Nuclear Cyclin D1 in MCF-7 Breast Cancer Cells: Discrimination between a Good and Bad Prognosis?

    doi: 10.1371/journal.pone.0042246

    Figure Lengend Snippet: Cyclin D1 is not required for p14ARF-induced cell growth (size, [forward scatter; FSC-A]). MCF7 cells were transfected in triplicate with siRNA as described in Figure 4 , and cells were harvested at day 3 and analysed using a BD LSRII cytometer. Flow cytometry data (20,000 events) were acquired using DIVA software (version 6.1.3), analysed with CellQuest Pro (version 6) and composite figures were generated with CANVAS (version X) software. A. To eliminate cell doublets and clumps from the analysis, single cells were gated (R1) from FSC-A versus FCS-H dot plots. A representative plot only is shown from mock-transfected cells. FSC-A histograms of the R1 populations were then used to indicate relative cell size of MCF-7 cells ± IPTG treatment after siRNA knockdown of cyclin D1, p53 and p21. Black line shows cells without IPTG and the red line shows cells treated with IPTG. B. Mean Fluorescent Intensity (MFI) analysis of FSC-A from triplicate biological experiments; data shown are mean MFI ± SD. C. Cell counts were performed using a haemocytometer. Data were presented as mean cell counts (x10 4 ) ± SD performed on three separate occasions.

    Article Snippet: Quantitative Real-time PCR (RT-QPCR) For gene expression changes, MCF-7 p14ARF cells were seeded at 1.0×105 cells/ml in 10% FCS DMEM for 24 h. Following 12h IPTG (5 mM) (Promega), ICI 182780 (10 nM), or vehicle treatment cells were homogenised in TRIzol (Invitrogen) and RNA extracted according to manufacturer’s instructions.

    Techniques: Transfection, Cytometry, Flow Cytometry, Software, Generated

    Cell cycle recurrence is abrogated by anti-estrogen ICI 182780. Cells were seeded at 200 cells per well and treated with combinations of IPTG, ICI 182780, E2 and vehicle over a period of 4 and 12 weeks and colonies were visualised with methylene blue staining. A. Colonies (+) were counted using a Bio-Rad ChemiDoc XRS + imager in biological triplicate. B. Representative wells showing IPTG and ICI 182780 and vehicle (control) treated colonies at week 4. C. IPTG treated cells at week 12.

    Journal: PLoS ONE

    Article Title: p14ARF Post-Transcriptional Regulation of Nuclear Cyclin D1 in MCF-7 Breast Cancer Cells: Discrimination between a Good and Bad Prognosis?

    doi: 10.1371/journal.pone.0042246

    Figure Lengend Snippet: Cell cycle recurrence is abrogated by anti-estrogen ICI 182780. Cells were seeded at 200 cells per well and treated with combinations of IPTG, ICI 182780, E2 and vehicle over a period of 4 and 12 weeks and colonies were visualised with methylene blue staining. A. Colonies (+) were counted using a Bio-Rad ChemiDoc XRS + imager in biological triplicate. B. Representative wells showing IPTG and ICI 182780 and vehicle (control) treated colonies at week 4. C. IPTG treated cells at week 12.

    Article Snippet: Quantitative Real-time PCR (RT-QPCR) For gene expression changes, MCF-7 p14ARF cells were seeded at 1.0×105 cells/ml in 10% FCS DMEM for 24 h. Following 12h IPTG (5 mM) (Promega), ICI 182780 (10 nM), or vehicle treatment cells were homogenised in TRIzol (Invitrogen) and RNA extracted according to manufacturer’s instructions.

    Techniques: Staining

    Regulation of cyclin D1, p53, and p21 mRNA and protein expression of in MCF-7p14ARF. Cells were treated with IPTG, ICI 182780 (ICI), or E2 for 12 h. Top panel: relative gene expression of A. CCDN1 , B. TP53 and C. CDKN1A mRNA quantified by qRT-PCR and relative expression normalised to control ACTB. Columns, Data represents the mean ± SEM of three separate experiments (** P

    Journal: PLoS ONE

    Article Title: p14ARF Post-Transcriptional Regulation of Nuclear Cyclin D1 in MCF-7 Breast Cancer Cells: Discrimination between a Good and Bad Prognosis?

    doi: 10.1371/journal.pone.0042246

    Figure Lengend Snippet: Regulation of cyclin D1, p53, and p21 mRNA and protein expression of in MCF-7p14ARF. Cells were treated with IPTG, ICI 182780 (ICI), or E2 for 12 h. Top panel: relative gene expression of A. CCDN1 , B. TP53 and C. CDKN1A mRNA quantified by qRT-PCR and relative expression normalised to control ACTB. Columns, Data represents the mean ± SEM of three separate experiments (** P

    Article Snippet: Quantitative Real-time PCR (RT-QPCR) For gene expression changes, MCF-7 p14ARF cells were seeded at 1.0×105 cells/ml in 10% FCS DMEM for 24 h. Following 12h IPTG (5 mM) (Promega), ICI 182780 (10 nM), or vehicle treatment cells were homogenised in TRIzol (Invitrogen) and RNA extracted according to manufacturer’s instructions.

    Techniques: Expressing, Quantitative RT-PCR

    3F3A mAb, but not anti-Gp78C, selectively recognizes dephosphorylated Gp78. (A) Cos7 cells were untransfected (Cos7 NT) or transfected with empty vector (PC3 EV), FLAG-tagged wild type (WT), S538A, phosphomimetic S538D or S538E, D536A mutant, or amino acid 533–541 deletion mutant (Δ3F3A) of Gp78. Cell lysates were immunoprecipitated with FLAG beads and probed with 3F3A, Gp78C, or FLAG mAbs. (B) Bacteria transformed with GST-tagged empty vector (GST-EV) or Gp78 C-terminal (GST-Gp78C) wild type (WT), S538A, S538D, S538E mutants, or amino acid 533–541 deletion mutant (Δ3F3A) were uninduced or induced by IPTG. Total bacterial lysates were dot blotted onto membranes and probed with 3F3A, Gp78C, or GST mAbs. (C) FLAG-tagged wild type of Gp78 (FLAG-Gp78-WT) or phosphorylation dominant-negative mutant (FLAG-Gp78 S538A) from Cos7 cells were immunoprecipitated and treated or not with CIAP and probed with 3F3A mAb, Gp78C, or FLAG mAb (±SEM; n = 3).

    Journal: Molecular Biology of the Cell

    Article Title: p38 MAP kinase–dependent phosphorylation of the Gp78 E3 ubiquitin ligase controls ER–mitochondria association and mitochondria motility

    doi: 10.1091/mbc.E15-02-0120

    Figure Lengend Snippet: 3F3A mAb, but not anti-Gp78C, selectively recognizes dephosphorylated Gp78. (A) Cos7 cells were untransfected (Cos7 NT) or transfected with empty vector (PC3 EV), FLAG-tagged wild type (WT), S538A, phosphomimetic S538D or S538E, D536A mutant, or amino acid 533–541 deletion mutant (Δ3F3A) of Gp78. Cell lysates were immunoprecipitated with FLAG beads and probed with 3F3A, Gp78C, or FLAG mAbs. (B) Bacteria transformed with GST-tagged empty vector (GST-EV) or Gp78 C-terminal (GST-Gp78C) wild type (WT), S538A, S538D, S538E mutants, or amino acid 533–541 deletion mutant (Δ3F3A) were uninduced or induced by IPTG. Total bacterial lysates were dot blotted onto membranes and probed with 3F3A, Gp78C, or GST mAbs. (C) FLAG-tagged wild type of Gp78 (FLAG-Gp78-WT) or phosphorylation dominant-negative mutant (FLAG-Gp78 S538A) from Cos7 cells were immunoprecipitated and treated or not with CIAP and probed with 3F3A mAb, Gp78C, or FLAG mAb (±SEM; n = 3).

    Article Snippet: GST-Gp78C protein expression was induced by 1 mM isopropyl-β-d -thiogalactoside (IPTG) at 37°C for 4 h. pE-SUMO His-Gp78 was purchased from LifeSensors and the His-hGp78C induced with 0.4 mM IPTG at 16°C overnight and purified with His60 Ni Superflow Resin (Clontech). pcDNA3-Flag dominant-negative MKK6(K82A) and constitutively active MKK6 (Glu) were gifts from Roger Davis (University of Massachusetts, Worcester, MA; Addgene plasmids 13519 and 13518; ).

    Techniques: Transfection, Plasmid Preparation, Mutagenesis, Immunoprecipitation, Transformation Assay, Dominant Negative Mutation

    Fig. 2. Electrophoretic mobility and purification of HEβ. ( A ) Western blot developed with anti-E-tag mAb–POD of whole-cell protein extracts from IPTG-induced E.coli UT5600 harboring pHEβ. Before loading onto the 10% polyacrylamide gel, the samples were resuspended in denaturing SDS–PAGE sample buffer and heated for 10 min at the indicated temperatures (25, 42, 50, 65 and 100°C). The faster mobility band of HEβ (f) corresponds to the folded conformation of the hybrid. When unfolded, HEβ migrates as a slower mobility band (u). ( B ) A sample of purified HEβ is shown after Coomassie Blue staining of a 10% SDS–polyacrylamide gel. ( C ) Western blot developed with anti- E-tag mAb–POD of purified HEβ samples treated at 25 (lane 1) or 100°C (lane 2) for 10 min before loading onto a 10% SDS– polyacrylamide gel. ( D ) The possible presence of contaminating OmpF porin in the purified HEβ was evaluated by western blot developed with a polyclonal serum against trimeric OmpF. Excess purified HEβ (10 µg, lanes 1 and 2) and a sample of purified OmpF (0.1 µg, lanes 3 and 4) as a control were loaded onto a 10% SDS–polyacrylamide gel after heating at 25 (lanes 1 and 3) or 100°C (lanes 2 and 4) for 10 min. Only the trimeric OmpF control (lane 3) was detected, ruling out the presence of OmpF in the purified HEβ sample.

    Journal: The EMBO Journal

    Article Title: Export of autotransported proteins proceeds through an oligomeric ring shaped byC-terminal domains

    doi:

    Figure Lengend Snippet: Fig. 2. Electrophoretic mobility and purification of HEβ. ( A ) Western blot developed with anti-E-tag mAb–POD of whole-cell protein extracts from IPTG-induced E.coli UT5600 harboring pHEβ. Before loading onto the 10% polyacrylamide gel, the samples were resuspended in denaturing SDS–PAGE sample buffer and heated for 10 min at the indicated temperatures (25, 42, 50, 65 and 100°C). The faster mobility band of HEβ (f) corresponds to the folded conformation of the hybrid. When unfolded, HEβ migrates as a slower mobility band (u). ( B ) A sample of purified HEβ is shown after Coomassie Blue staining of a 10% SDS–polyacrylamide gel. ( C ) Western blot developed with anti- E-tag mAb–POD of purified HEβ samples treated at 25 (lane 1) or 100°C (lane 2) for 10 min before loading onto a 10% SDS– polyacrylamide gel. ( D ) The possible presence of contaminating OmpF porin in the purified HEβ was evaluated by western blot developed with a polyclonal serum against trimeric OmpF. Excess purified HEβ (10 µg, lanes 1 and 2) and a sample of purified OmpF (0.1 µg, lanes 3 and 4) as a control were loaded onto a 10% SDS–polyacrylamide gel after heating at 25 (lanes 1 and 3) or 100°C (lanes 2 and 4) for 10 min. Only the trimeric OmpF control (lane 3) was detected, ruling out the presence of OmpF in the purified HEβ sample.

    Article Snippet: Escherichia coli UT5600 (pHEβ) cells, grown in LB + Cm at 30°C, were harvested after 3 h induction with 0.1 mM IPTG (final OD600 ∼1.0) and resuspended in one-tenth of the original culture volume of PBS containing 0.2 mM DSP (Pierce).

    Techniques: Purification, Western Blot, SDS Page, Staining