full length trbp1 orf Search Results


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  • 88
    TaKaRa ptre2pur
    TRBP overexpression protects cells from arsenite-induced apoptosis. ( A ) Establishment of a stable doxycycline-inducible HeLa cell line overexpressing flag-TRBP. HeLa-Tet off (Clontech) cells were transfected with either <t>FlagTRBP/pTRE2pur</t> expression construct or pTRE2pur (Clontech) empty vector (EV). Puromycin resistant clones were isolated, characterized, and one clone (TRBP-HeLa) that showed inducible expression of FlagTRBP was selected for further studies. Induction of FlagTRBP expression after removal of doxycycline from the growth medium at indicated time points is shown. Western blot analysis of cell lysates using 50 μg of total protein was performed using anti-Flag and anti-β-actin antibodies. The black line between lanes 3 and 4 represents where different lanes from the same western blot were joined. ( B ) DNA Fragmentation Analysis. TRBP-HeLa cells overexpressing FlagTRBP (Lanes 1–4) or EV-HeLa control cells (Lanes 5–8) were treated with 10 μM sodium arsenite for 48, 72, and 96 hours, fragmented DNA was isolated and analyzed. M: 100-bp ladder, Lanes 1 5: 48 hr treatment. Lanes 2 6: 72 hr treatment, Lanes 3 7: 96 hr treatment, and Lanes 4 8: untreated cells. ( C ) Analysis of PARP cleavage in response to arsenite treatment. TRBP-HeLa and EV-HeLa cells were treated with 25 μM sodium arsenite for the indicated time points, and western blot analysis using anti-PARP antibody was performed on cell lysates containing 50 μg of total protein to assess increases in poly ADP-ribose polymerase (PARP1) cleavage. Western blot was also performed with anti-GAPDH antibody to ensure equal protein in all samples. ( D ) Quantification of PARP cleavage. PARP1 and cleaved PARP1 bands were quantified using ImageQuant TL Software. The percentage of cleaved PARP1 was calculated as (cleaved PARP1 band intensity/cleaved + uncleaved bands intensities) X 100. Bars represent percentages of cleaved PARP1 from 3 independent experiments. Error bars represent standard deviation (S.D.) from three experiments. Student’s t tests were performed to determine statistical significance – ns: not significant, asterisk: p value of 0.000352.
    Ptre2pur, supplied by TaKaRa, used in various techniques. Bioz Stars score: 88/100, based on 95 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa ecorv restriction sites
    TRBP overexpression protects cells from arsenite-induced apoptosis. ( A ) Establishment of a stable doxycycline-inducible HeLa cell line overexpressing flag-TRBP. HeLa-Tet off (Clontech) cells were transfected with either <t>FlagTRBP/pTRE2pur</t> expression construct or pTRE2pur (Clontech) empty vector (EV). Puromycin resistant clones were isolated, characterized, and one clone (TRBP-HeLa) that showed inducible expression of FlagTRBP was selected for further studies. Induction of FlagTRBP expression after removal of doxycycline from the growth medium at indicated time points is shown. Western blot analysis of cell lysates using 50 μg of total protein was performed using anti-Flag and anti-β-actin antibodies. The black line between lanes 3 and 4 represents where different lanes from the same western blot were joined. ( B ) DNA Fragmentation Analysis. TRBP-HeLa cells overexpressing FlagTRBP (Lanes 1–4) or EV-HeLa control cells (Lanes 5–8) were treated with 10 μM sodium arsenite for 48, 72, and 96 hours, fragmented DNA was isolated and analyzed. M: 100-bp ladder, Lanes 1 5: 48 hr treatment. Lanes 2 6: 72 hr treatment, Lanes 3 7: 96 hr treatment, and Lanes 4 8: untreated cells. ( C ) Analysis of PARP cleavage in response to arsenite treatment. TRBP-HeLa and EV-HeLa cells were treated with 25 μM sodium arsenite for the indicated time points, and western blot analysis using anti-PARP antibody was performed on cell lysates containing 50 μg of total protein to assess increases in poly ADP-ribose polymerase (PARP1) cleavage. Western blot was also performed with anti-GAPDH antibody to ensure equal protein in all samples. ( D ) Quantification of PARP cleavage. PARP1 and cleaved PARP1 bands were quantified using ImageQuant TL Software. The percentage of cleaved PARP1 was calculated as (cleaved PARP1 band intensity/cleaved + uncleaved bands intensities) X 100. Bars represent percentages of cleaved PARP1 from 3 independent experiments. Error bars represent standard deviation (S.D.) from three experiments. Student’s t tests were performed to determine statistical significance – ns: not significant, asterisk: p value of 0.000352.
    Ecorv Restriction Sites, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ecorv restriction sites - by Bioz Stars, 2020-08
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    89
    TaKaRa tetracycline responsive vector
    TRBP overexpression protects cells from arsenite-induced apoptosis. ( A ) Establishment of a stable doxycycline-inducible HeLa cell line overexpressing flag-TRBP. HeLa-Tet off (Clontech) cells were transfected with either <t>FlagTRBP/pTRE2pur</t> expression construct or pTRE2pur (Clontech) empty vector (EV). Puromycin resistant clones were isolated, characterized, and one clone (TRBP-HeLa) that showed inducible expression of FlagTRBP was selected for further studies. Induction of FlagTRBP expression after removal of doxycycline from the growth medium at indicated time points is shown. Western blot analysis of cell lysates using 50 μg of total protein was performed using anti-Flag and anti-β-actin antibodies. The black line between lanes 3 and 4 represents where different lanes from the same western blot were joined. ( B ) DNA Fragmentation Analysis. TRBP-HeLa cells overexpressing FlagTRBP (Lanes 1–4) or EV-HeLa control cells (Lanes 5–8) were treated with 10 μM sodium arsenite for 48, 72, and 96 hours, fragmented DNA was isolated and analyzed. M: 100-bp ladder, Lanes 1 5: 48 hr treatment. Lanes 2 6: 72 hr treatment, Lanes 3 7: 96 hr treatment, and Lanes 4 8: untreated cells. ( C ) Analysis of PARP cleavage in response to arsenite treatment. TRBP-HeLa and EV-HeLa cells were treated with 25 μM sodium arsenite for the indicated time points, and western blot analysis using anti-PARP antibody was performed on cell lysates containing 50 μg of total protein to assess increases in poly ADP-ribose polymerase (PARP1) cleavage. Western blot was also performed with anti-GAPDH antibody to ensure equal protein in all samples. ( D ) Quantification of PARP cleavage. PARP1 and cleaved PARP1 bands were quantified using ImageQuant TL Software. The percentage of cleaved PARP1 was calculated as (cleaved PARP1 band intensity/cleaved + uncleaved bands intensities) X 100. Bars represent percentages of cleaved PARP1 from 3 independent experiments. Error bars represent standard deviation (S.D.) from three experiments. Student’s t tests were performed to determine statistical significance – ns: not significant, asterisk: p value of 0.000352.
    Tetracycline Responsive Vector, supplied by TaKaRa, used in various techniques. Bioz Stars score: 89/100, based on 72 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Promega pgemt easy vector
    TRBP overexpression protects cells from arsenite-induced apoptosis. ( A ) Establishment of a stable doxycycline-inducible HeLa cell line overexpressing flag-TRBP. HeLa-Tet off (Clontech) cells were transfected with either <t>FlagTRBP/pTRE2pur</t> expression construct or pTRE2pur (Clontech) empty vector (EV). Puromycin resistant clones were isolated, characterized, and one clone (TRBP-HeLa) that showed inducible expression of FlagTRBP was selected for further studies. Induction of FlagTRBP expression after removal of doxycycline from the growth medium at indicated time points is shown. Western blot analysis of cell lysates using 50 μg of total protein was performed using anti-Flag and anti-β-actin antibodies. The black line between lanes 3 and 4 represents where different lanes from the same western blot were joined. ( B ) DNA Fragmentation Analysis. TRBP-HeLa cells overexpressing FlagTRBP (Lanes 1–4) or EV-HeLa control cells (Lanes 5–8) were treated with 10 μM sodium arsenite for 48, 72, and 96 hours, fragmented DNA was isolated and analyzed. M: 100-bp ladder, Lanes 1 5: 48 hr treatment. Lanes 2 6: 72 hr treatment, Lanes 3 7: 96 hr treatment, and Lanes 4 8: untreated cells. ( C ) Analysis of PARP cleavage in response to arsenite treatment. TRBP-HeLa and EV-HeLa cells were treated with 25 μM sodium arsenite for the indicated time points, and western blot analysis using anti-PARP antibody was performed on cell lysates containing 50 μg of total protein to assess increases in poly ADP-ribose polymerase (PARP1) cleavage. Western blot was also performed with anti-GAPDH antibody to ensure equal protein in all samples. ( D ) Quantification of PARP cleavage. PARP1 and cleaved PARP1 bands were quantified using ImageQuant TL Software. The percentage of cleaved PARP1 was calculated as (cleaved PARP1 band intensity/cleaved + uncleaved bands intensities) X 100. Bars represent percentages of cleaved PARP1 from 3 independent experiments. Error bars represent standard deviation (S.D.) from three experiments. Student’s t tests were performed to determine statistical significance – ns: not significant, asterisk: p value of 0.000352.
    Pgemt Easy Vector, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 6738 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    TRBP overexpression protects cells from arsenite-induced apoptosis. ( A ) Establishment of a stable doxycycline-inducible HeLa cell line overexpressing flag-TRBP. HeLa-Tet off (Clontech) cells were transfected with either FlagTRBP/pTRE2pur expression construct or pTRE2pur (Clontech) empty vector (EV). Puromycin resistant clones were isolated, characterized, and one clone (TRBP-HeLa) that showed inducible expression of FlagTRBP was selected for further studies. Induction of FlagTRBP expression after removal of doxycycline from the growth medium at indicated time points is shown. Western blot analysis of cell lysates using 50 μg of total protein was performed using anti-Flag and anti-β-actin antibodies. The black line between lanes 3 and 4 represents where different lanes from the same western blot were joined. ( B ) DNA Fragmentation Analysis. TRBP-HeLa cells overexpressing FlagTRBP (Lanes 1–4) or EV-HeLa control cells (Lanes 5–8) were treated with 10 μM sodium arsenite for 48, 72, and 96 hours, fragmented DNA was isolated and analyzed. M: 100-bp ladder, Lanes 1 5: 48 hr treatment. Lanes 2 6: 72 hr treatment, Lanes 3 7: 96 hr treatment, and Lanes 4 8: untreated cells. ( C ) Analysis of PARP cleavage in response to arsenite treatment. TRBP-HeLa and EV-HeLa cells were treated with 25 μM sodium arsenite for the indicated time points, and western blot analysis using anti-PARP antibody was performed on cell lysates containing 50 μg of total protein to assess increases in poly ADP-ribose polymerase (PARP1) cleavage. Western blot was also performed with anti-GAPDH antibody to ensure equal protein in all samples. ( D ) Quantification of PARP cleavage. PARP1 and cleaved PARP1 bands were quantified using ImageQuant TL Software. The percentage of cleaved PARP1 was calculated as (cleaved PARP1 band intensity/cleaved + uncleaved bands intensities) X 100. Bars represent percentages of cleaved PARP1 from 3 independent experiments. Error bars represent standard deviation (S.D.) from three experiments. Student’s t tests were performed to determine statistical significance – ns: not significant, asterisk: p value of 0.000352.

    Journal: Scientific Reports

    Article Title: Stress-induced TRBP phosphorylation enhances its interaction with PKR to regulate cellular survival

    doi: 10.1038/s41598-018-19360-8

    Figure Lengend Snippet: TRBP overexpression protects cells from arsenite-induced apoptosis. ( A ) Establishment of a stable doxycycline-inducible HeLa cell line overexpressing flag-TRBP. HeLa-Tet off (Clontech) cells were transfected with either FlagTRBP/pTRE2pur expression construct or pTRE2pur (Clontech) empty vector (EV). Puromycin resistant clones were isolated, characterized, and one clone (TRBP-HeLa) that showed inducible expression of FlagTRBP was selected for further studies. Induction of FlagTRBP expression after removal of doxycycline from the growth medium at indicated time points is shown. Western blot analysis of cell lysates using 50 μg of total protein was performed using anti-Flag and anti-β-actin antibodies. The black line between lanes 3 and 4 represents where different lanes from the same western blot were joined. ( B ) DNA Fragmentation Analysis. TRBP-HeLa cells overexpressing FlagTRBP (Lanes 1–4) or EV-HeLa control cells (Lanes 5–8) were treated with 10 μM sodium arsenite for 48, 72, and 96 hours, fragmented DNA was isolated and analyzed. M: 100-bp ladder, Lanes 1 5: 48 hr treatment. Lanes 2 6: 72 hr treatment, Lanes 3 7: 96 hr treatment, and Lanes 4 8: untreated cells. ( C ) Analysis of PARP cleavage in response to arsenite treatment. TRBP-HeLa and EV-HeLa cells were treated with 25 μM sodium arsenite for the indicated time points, and western blot analysis using anti-PARP antibody was performed on cell lysates containing 50 μg of total protein to assess increases in poly ADP-ribose polymerase (PARP1) cleavage. Western blot was also performed with anti-GAPDH antibody to ensure equal protein in all samples. ( D ) Quantification of PARP cleavage. PARP1 and cleaved PARP1 bands were quantified using ImageQuant TL Software. The percentage of cleaved PARP1 was calculated as (cleaved PARP1 band intensity/cleaved + uncleaved bands intensities) X 100. Bars represent percentages of cleaved PARP1 from 3 independent experiments. Error bars represent standard deviation (S.D.) from three experiments. Student’s t tests were performed to determine statistical significance – ns: not significant, asterisk: p value of 0.000352.

    Article Snippet: Full length TRBP1 ORF with an N-terminal Flag tag from Flag TRBP/BSIIKS+ was inserted into the NotI and EcoRV restriction sites of the tetracycline-responsive vector, pTRE2pur (Clontech) to generate Flag TRBP/pTRE2pur.

    Techniques: Over Expression, Transfection, Expressing, Construct, Plasmid Preparation, Clone Assay, Isolation, Western Blot, Software, Standard Deviation