control gene fopflash Search Results


fopflash constructs  (Addgene inc)
96
Addgene inc fopflash constructs
USP2a upregulates β-catenin protein and promotes its transcriptional activity. A. Each SFB-tagged deubiquitinase was co-transfected with MYC-tagged β-catenin into HEK293T cells, followed by pulldown with S-protein beads and immunoblotting with antibodies against FLAG and MYC. B. Either β-catenin-responsive TOPflash or its mutant <t>FOPflash</t> construct was co-transfected with each SFB-tagged deubiquitinase and Renilla luciferase into HEK293T cells. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity. MYC-tagged β-catenin serves as a positive control. Error bars are S.D. C. SFB-tagged GFP, USP2a, USP26, and USP42 were co-transfected with MYC-tagged β-catenin into HEK293T cells, and pulled down with S-protein beads. Antibodies against MYC and FLAG were used to detect β-catenin and DUBs. CypB (cyclophilin B) serves as the loading control. D. Left panel: luciferase reporter assay validating that the 4 candidate deubiquitinases (USP2a, USP36, DUB3, and OTUD7B) promote the transcriptional activity of β-catenin. Error bars are S.D. ***: P < 0.001. Right panel: immunoblotting of HSP90 and SFB-tagged GFP, USP2a, USP36, DUB3, and OTUD7B in HEK293T cells. HSP90 serves as the loading control. T: TOPflash; F: FOPflash. E. Each SFB-tagged candidate deubiquitinase was co-transfected with <t>HA-tagged</t> <t>ubiquitin</t> and MYC-tagged β-catenin into HEK293T cells. After MG132 treatment for 6 hours, β-catenin was immunoprecipitated with a MYC-specific antibody, followed by immunoblotting with antibodies against HA and MYC. F. Immunoblotting of β-catenin, FLAG, and HSP90 in HEK293T cells transfected with SFB-tagged GFP, USP2a, USP36, DUB3, or OTUD7B. G. qPCR of CTNNB1 (the gene that encodes β-catenin) in HEK293T cells transfected with the empty vector, wild-type USP2a, or the catalytically inactive mutant (C276A). Error bars are S.D. n.s.: not significant. H. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNAs. I. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNA with or without ectopic expression of USP2a.
Fopflash Constructs, supplied by Addgene inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/control+gene+fopflash/pRK5-HA-Ubiquitin-WT+(Plasmid+%2317608)/pmc06176183-47-3-7
Average 96 stars, based on 1 article reviews
fopflash constructs - by Bioz Stars, 2026-09
96/100 stars
  Buy from Supplier
control fopflash reporter plasmid dna  (Addgene inc)
94
Addgene inc control fopflash reporter plasmid dna
USP2a upregulates β-catenin protein and promotes its transcriptional activity. A. Each SFB-tagged deubiquitinase was co-transfected with MYC-tagged β-catenin into HEK293T cells, followed by pulldown with S-protein beads and immunoblotting with antibodies against FLAG and MYC. B. Either β-catenin-responsive TOPflash or its mutant <t>FOPflash</t> construct was co-transfected with each SFB-tagged deubiquitinase and Renilla luciferase into HEK293T cells. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity. MYC-tagged β-catenin serves as a positive control. Error bars are S.D. C. SFB-tagged GFP, USP2a, USP26, and USP42 were co-transfected with MYC-tagged β-catenin into HEK293T cells, and pulled down with S-protein beads. Antibodies against MYC and FLAG were used to detect β-catenin and DUBs. CypB (cyclophilin B) serves as the loading control. D. Left panel: luciferase reporter assay validating that the 4 candidate deubiquitinases (USP2a, USP36, DUB3, and OTUD7B) promote the transcriptional activity of β-catenin. Error bars are S.D. ***: P < 0.001. Right panel: immunoblotting of HSP90 and SFB-tagged GFP, USP2a, USP36, DUB3, and OTUD7B in HEK293T cells. HSP90 serves as the loading control. T: TOPflash; F: FOPflash. E. Each SFB-tagged candidate deubiquitinase was co-transfected with <t>HA-tagged</t> <t>ubiquitin</t> and MYC-tagged β-catenin into HEK293T cells. After MG132 treatment for 6 hours, β-catenin was immunoprecipitated with a MYC-specific antibody, followed by immunoblotting with antibodies against HA and MYC. F. Immunoblotting of β-catenin, FLAG, and HSP90 in HEK293T cells transfected with SFB-tagged GFP, USP2a, USP36, DUB3, or OTUD7B. G. qPCR of CTNNB1 (the gene that encodes β-catenin) in HEK293T cells transfected with the empty vector, wild-type USP2a, or the catalytically inactive mutant (C276A). Error bars are S.D. n.s.: not significant. H. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNAs. I. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNA with or without ectopic expression of USP2a.
Control Fopflash Reporter Plasmid Dna, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/control+gene+fopflash/M51+Super+8x+FOPFlash+(TOPFlash+mutant)+(Plasmid+%2312457)/10__1530_slash_erc___12___0328-68-35-51
Average 94 stars, based on 1 article reviews
control fopflash reporter plasmid dna - by Bioz Stars, 2026-09
94/100 stars
  Buy from Supplier

Image Search Results


USP2a upregulates β-catenin protein and promotes its transcriptional activity. A. Each SFB-tagged deubiquitinase was co-transfected with MYC-tagged β-catenin into HEK293T cells, followed by pulldown with S-protein beads and immunoblotting with antibodies against FLAG and MYC. B. Either β-catenin-responsive TOPflash or its mutant FOPflash construct was co-transfected with each SFB-tagged deubiquitinase and Renilla luciferase into HEK293T cells. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity. MYC-tagged β-catenin serves as a positive control. Error bars are S.D. C. SFB-tagged GFP, USP2a, USP26, and USP42 were co-transfected with MYC-tagged β-catenin into HEK293T cells, and pulled down with S-protein beads. Antibodies against MYC and FLAG were used to detect β-catenin and DUBs. CypB (cyclophilin B) serves as the loading control. D. Left panel: luciferase reporter assay validating that the 4 candidate deubiquitinases (USP2a, USP36, DUB3, and OTUD7B) promote the transcriptional activity of β-catenin. Error bars are S.D. ***: P < 0.001. Right panel: immunoblotting of HSP90 and SFB-tagged GFP, USP2a, USP36, DUB3, and OTUD7B in HEK293T cells. HSP90 serves as the loading control. T: TOPflash; F: FOPflash. E. Each SFB-tagged candidate deubiquitinase was co-transfected with HA-tagged ubiquitin and MYC-tagged β-catenin into HEK293T cells. After MG132 treatment for 6 hours, β-catenin was immunoprecipitated with a MYC-specific antibody, followed by immunoblotting with antibodies against HA and MYC. F. Immunoblotting of β-catenin, FLAG, and HSP90 in HEK293T cells transfected with SFB-tagged GFP, USP2a, USP36, DUB3, or OTUD7B. G. qPCR of CTNNB1 (the gene that encodes β-catenin) in HEK293T cells transfected with the empty vector, wild-type USP2a, or the catalytically inactive mutant (C276A). Error bars are S.D. n.s.: not significant. H. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNAs. I. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNA with or without ectopic expression of USP2a.

Journal: American Journal of Cancer Research

Article Title: Ubiquitin-specific peptidase 2a (USP2a) deubiquitinates and stabilizes β-catenin

doi:

Figure Lengend Snippet: USP2a upregulates β-catenin protein and promotes its transcriptional activity. A. Each SFB-tagged deubiquitinase was co-transfected with MYC-tagged β-catenin into HEK293T cells, followed by pulldown with S-protein beads and immunoblotting with antibodies against FLAG and MYC. B. Either β-catenin-responsive TOPflash or its mutant FOPflash construct was co-transfected with each SFB-tagged deubiquitinase and Renilla luciferase into HEK293T cells. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity. MYC-tagged β-catenin serves as a positive control. Error bars are S.D. C. SFB-tagged GFP, USP2a, USP26, and USP42 were co-transfected with MYC-tagged β-catenin into HEK293T cells, and pulled down with S-protein beads. Antibodies against MYC and FLAG were used to detect β-catenin and DUBs. CypB (cyclophilin B) serves as the loading control. D. Left panel: luciferase reporter assay validating that the 4 candidate deubiquitinases (USP2a, USP36, DUB3, and OTUD7B) promote the transcriptional activity of β-catenin. Error bars are S.D. ***: P < 0.001. Right panel: immunoblotting of HSP90 and SFB-tagged GFP, USP2a, USP36, DUB3, and OTUD7B in HEK293T cells. HSP90 serves as the loading control. T: TOPflash; F: FOPflash. E. Each SFB-tagged candidate deubiquitinase was co-transfected with HA-tagged ubiquitin and MYC-tagged β-catenin into HEK293T cells. After MG132 treatment for 6 hours, β-catenin was immunoprecipitated with a MYC-specific antibody, followed by immunoblotting with antibodies against HA and MYC. F. Immunoblotting of β-catenin, FLAG, and HSP90 in HEK293T cells transfected with SFB-tagged GFP, USP2a, USP36, DUB3, or OTUD7B. G. qPCR of CTNNB1 (the gene that encodes β-catenin) in HEK293T cells transfected with the empty vector, wild-type USP2a, or the catalytically inactive mutant (C276A). Error bars are S.D. n.s.: not significant. H. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNAs. I. Immunoblotting of USP2a, β-catenin, and CypB in BT549 cells transduced with USP2 shRNA with or without ectopic expression of USP2a.

Article Snippet: HA-ubiquitin, TOPflash, and FOPflash constructs were from Addgene (plasmid number: 17608, 12456, and 12457).

Techniques: Activity Assay, Transfection, Western Blot, Mutagenesis, Construct, Luciferase, Positive Control, Control, Reporter Assay, Ubiquitin Proteomics, Immunoprecipitation, Plasmid Preparation, Transduction, shRNA, Expressing

USP2a activates Wnt/β-catenin signaling. A. Immunoblotting of FLAG-USP2a, β-catenin, HSP90 (cytoplasmic marker), and Lamin B1 (nuclear marker) in cytoplasmic and nuclear fractions of HEK293T cells transfected with the empty vector or SFB-USP2a. B. Top panel: either β-catenin-responsive TOPflash or its mutant FOPflash construct was co-transfected with SFB-tagged GFP, USP2a, or USP2aC276A and Renilla luciferase into HEK293T cells. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity. Error bars are S.D. **: P < 0.01. Bottom panel: immunoblotting of SFB-USP2a, SFB-USP2aC276A, and CypB in HEK293T cells. T: TOPflash; F: FOPflash. C. Top panel: either β-catenin-responsive TOPflash or its mutant FOPflash construct was co-transfected with the SFB vector or SFB-USP2a and Renilla luciferase into HEK293T cells. Cells were treated with DMSO or 3 µM ML364 for 24 hours. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity, and then TOPflash reads were normalized to FOPflash reads. Error bars are S.E.M. *: P < 0.05; **: P < 0.01. Bottom panel: immunoblotting of SFB-USP2a and CypB in HEK293T cells. D: DMSO; M: ML364. D. Human Wnt signaling targets PCR array analysis of HEK293T cells transfected with SFB-USP2a. Gene expression levels in SFB-USP2a-transfected cells were compared to those in empty vector-transfected cells (log2 scale). E. qPCR of BIRC5, CUBN, FGF7, CDON, BTRC, ANTXR1, and IGF2 in HEK293T cells transfected with the empty vector, SFB-USP2a, or SFB-USP2aC276A. Error bars are S.E.M. *: P < 0.05; **: P < 0.01; ***: P < 0.001. F. qPCR of BIRC5, CUBN, FGF7, CDON, BTRC, ANTXR1, and IGF2 in BT549 cells transduced with USP2 shRNAs or a scramble control (Scr). Error bars are S.E.M. *: P < 0.05; **: P < 0.01; ***: P < 0.001; #: not detectable.

Journal: American Journal of Cancer Research

Article Title: Ubiquitin-specific peptidase 2a (USP2a) deubiquitinates and stabilizes β-catenin

doi:

Figure Lengend Snippet: USP2a activates Wnt/β-catenin signaling. A. Immunoblotting of FLAG-USP2a, β-catenin, HSP90 (cytoplasmic marker), and Lamin B1 (nuclear marker) in cytoplasmic and nuclear fractions of HEK293T cells transfected with the empty vector or SFB-USP2a. B. Top panel: either β-catenin-responsive TOPflash or its mutant FOPflash construct was co-transfected with SFB-tagged GFP, USP2a, or USP2aC276A and Renilla luciferase into HEK293T cells. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity. Error bars are S.D. **: P < 0.01. Bottom panel: immunoblotting of SFB-USP2a, SFB-USP2aC276A, and CypB in HEK293T cells. T: TOPflash; F: FOPflash. C. Top panel: either β-catenin-responsive TOPflash or its mutant FOPflash construct was co-transfected with the SFB vector or SFB-USP2a and Renilla luciferase into HEK293T cells. Cells were treated with DMSO or 3 µM ML364 for 24 hours. A dual luciferase assay was used to determine β-catenin activity. Firefly luciferase activity was normalized to Renilla luciferase activity, and then TOPflash reads were normalized to FOPflash reads. Error bars are S.E.M. *: P < 0.05; **: P < 0.01. Bottom panel: immunoblotting of SFB-USP2a and CypB in HEK293T cells. D: DMSO; M: ML364. D. Human Wnt signaling targets PCR array analysis of HEK293T cells transfected with SFB-USP2a. Gene expression levels in SFB-USP2a-transfected cells were compared to those in empty vector-transfected cells (log2 scale). E. qPCR of BIRC5, CUBN, FGF7, CDON, BTRC, ANTXR1, and IGF2 in HEK293T cells transfected with the empty vector, SFB-USP2a, or SFB-USP2aC276A. Error bars are S.E.M. *: P < 0.05; **: P < 0.01; ***: P < 0.001. F. qPCR of BIRC5, CUBN, FGF7, CDON, BTRC, ANTXR1, and IGF2 in BT549 cells transduced with USP2 shRNAs or a scramble control (Scr). Error bars are S.E.M. *: P < 0.05; **: P < 0.01; ***: P < 0.001; #: not detectable.

Article Snippet: HA-ubiquitin, TOPflash, and FOPflash constructs were from Addgene (plasmid number: 17608, 12456, and 12457).

Techniques: Western Blot, Marker, Transfection, Plasmid Preparation, Mutagenesis, Construct, Luciferase, Activity Assay, Gene Expression, Transduction, Control