transfact Search Results


transfact transfection reagent  (Promega)
90
Promega transfact transfection reagent
T3 blocks ras-mediated proliferation and cyclin D1 expression. (A) BrdU incorporation was determined as indicated in Materials and Methods in N2a-β cells cotransfected with EGFP and Ha-rasval12 and incubated in the presence or absence of 5 nM T3 for 48 h. (B) Representative Western blot of cyclin D1, p27Kip, and p21Cip in cells transfected with 50 ng of Ha-rasval12 or with the same amount of an empty vector. After <t>transfection</t> cells were treated for 16 h in the presence or absence of 5 nM T3. (C) Flow cytometry analysis of cells transfected with EGFP and Ha-rasval12 alone or in combination with an expression vector for cyclin D1. The number of cells in S+G2/M phases was determined in control cells and in cells incubated with T3, as indicated in Materials and Methods, and the percentage of inhibition by the hormone is shown for each experimental condition.
Transfact Transfection Reagent, supplied by Promega, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 90 stars, based on 1 article reviews
transfact transfection reagent - by Bioz Stars, 2026-03
90/100 stars
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transfact  (Promega)
90
Promega transfact
T3 blocks ras-mediated proliferation and cyclin D1 expression. (A) BrdU incorporation was determined as indicated in Materials and Methods in N2a-β cells cotransfected with EGFP and Ha-rasval12 and incubated in the presence or absence of 5 nM T3 for 48 h. (B) Representative Western blot of cyclin D1, p27Kip, and p21Cip in cells transfected with 50 ng of Ha-rasval12 or with the same amount of an empty vector. After <t>transfection</t> cells were treated for 16 h in the presence or absence of 5 nM T3. (C) Flow cytometry analysis of cells transfected with EGFP and Ha-rasval12 alone or in combination with an expression vector for cyclin D1. The number of cells in S+G2/M phases was determined in control cells and in cells incubated with T3, as indicated in Materials and Methods, and the percentage of inhibition by the hormone is shown for each experimental condition.
Transfact, supplied by Promega, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/transfact/product/Promega
Average 90 stars, based on 1 article reviews
transfact - by Bioz Stars, 2026-03
90/100 stars
  Buy from Supplier

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T3 blocks ras-mediated proliferation and cyclin D1 expression. (A) BrdU incorporation was determined as indicated in Materials and Methods in N2a-β cells cotransfected with EGFP and Ha-rasval12 and incubated in the presence or absence of 5 nM T3 for 48 h. (B) Representative Western blot of cyclin D1, p27Kip, and p21Cip in cells transfected with 50 ng of Ha-rasval12 or with the same amount of an empty vector. After transfection cells were treated for 16 h in the presence or absence of 5 nM T3. (C) Flow cytometry analysis of cells transfected with EGFP and Ha-rasval12 alone or in combination with an expression vector for cyclin D1. The number of cells in S+G2/M phases was determined in control cells and in cells incubated with T3, as indicated in Materials and Methods, and the percentage of inhibition by the hormone is shown for each experimental condition.

Journal:

Article Title: The Thyroid Hormone Receptor Is a Suppressor of ras -Mediated Transcription, Proliferation, and Transformation

doi: 10.1128/MCB.24.17.7514-7523.2004

Figure Lengend Snippet: T3 blocks ras-mediated proliferation and cyclin D1 expression. (A) BrdU incorporation was determined as indicated in Materials and Methods in N2a-β cells cotransfected with EGFP and Ha-rasval12 and incubated in the presence or absence of 5 nM T3 for 48 h. (B) Representative Western blot of cyclin D1, p27Kip, and p21Cip in cells transfected with 50 ng of Ha-rasval12 or with the same amount of an empty vector. After transfection cells were treated for 16 h in the presence or absence of 5 nM T3. (C) Flow cytometry analysis of cells transfected with EGFP and Ha-rasval12 alone or in combination with an expression vector for cyclin D1. The number of cells in S+G2/M phases was determined in control cells and in cells incubated with T3, as indicated in Materials and Methods, and the percentage of inhibition by the hormone is shown for each experimental condition.

Article Snippet: N2a-β cells were plated in 24-well plates and cotransfected by using Transfact transfection reagent (Promega) with 10 ng of a plasmid encoding enhanced green fluorescence protein (p-EGFP-C1; Clontech) and 40 ng of pCEFL-Ha- ras val12 or the corresponding empty vector.

Techniques: Expressing, BrdU Incorporation Assay, Incubation, Western Blot, Transfection, Plasmid Preparation, Flow Cytometry, Inhibition

T3 represses activation of the cyclin D1 promoter by oncogenic ras. (A) Transient-transfection assays with a reporter plasmid containing the cyclin D1 promoter (sequences −1720/+141) and 50 ng of Ha-rasval12 or the empty vector. The luciferase activity was determined in cells incubated for 36 h with the concentrations of T3 indicated and are expressed as the fold induction over the values obtained in the untreated cells transfected with the empty vector. (B) The reporter activity was determined after different periods of incubation in the presence or absence of 5 nM T3. (C) Cells were transfected with activated forms of Ha-, K-, and N-rasval12 (50 ng) and incubated with 5 nM T3 for 36 h before determination of luciferase activity. In the lower panels, the levels of Ras proteins were determined by Western blotting with a pan-Ras antibody, and the levels of Erk2 were determined as a loading control. (D) TRβ expression was determined by Western blotting with 20 μg of proteins from parental N2a, N2a-β, and pituitary GH4C1 cells. (E) Cyclin D1 promoter activity was determined in pituitary GH4C1 cells transiently transfected with 50 ng of Ha-rasval12 and treated for 36 h in the absence and presence of 5 nM T3.

Journal:

Article Title: The Thyroid Hormone Receptor Is a Suppressor of ras -Mediated Transcription, Proliferation, and Transformation

doi: 10.1128/MCB.24.17.7514-7523.2004

Figure Lengend Snippet: T3 represses activation of the cyclin D1 promoter by oncogenic ras. (A) Transient-transfection assays with a reporter plasmid containing the cyclin D1 promoter (sequences −1720/+141) and 50 ng of Ha-rasval12 or the empty vector. The luciferase activity was determined in cells incubated for 36 h with the concentrations of T3 indicated and are expressed as the fold induction over the values obtained in the untreated cells transfected with the empty vector. (B) The reporter activity was determined after different periods of incubation in the presence or absence of 5 nM T3. (C) Cells were transfected with activated forms of Ha-, K-, and N-rasval12 (50 ng) and incubated with 5 nM T3 for 36 h before determination of luciferase activity. In the lower panels, the levels of Ras proteins were determined by Western blotting with a pan-Ras antibody, and the levels of Erk2 were determined as a loading control. (D) TRβ expression was determined by Western blotting with 20 μg of proteins from parental N2a, N2a-β, and pituitary GH4C1 cells. (E) Cyclin D1 promoter activity was determined in pituitary GH4C1 cells transiently transfected with 50 ng of Ha-rasval12 and treated for 36 h in the absence and presence of 5 nM T3.

Article Snippet: N2a-β cells were plated in 24-well plates and cotransfected by using Transfact transfection reagent (Promega) with 10 ng of a plasmid encoding enhanced green fluorescence protein (p-EGFP-C1; Clontech) and 40 ng of pCEFL-Ha- ras val12 or the corresponding empty vector.

Techniques: Activation Assay, Transfection, Plasmid Preparation, Luciferase, Activity Assay, Incubation, Western Blot, Expressing

The Ras/Erk pathway is the main effector for Ras-mediated stimulation of the cyclin D1 promoter in neuroblastoma cells. (A) N2a-β cells were transfected with the cyclin D1 promoter construct and 50 ng of Ha-rasval12. Reporter activity was determined in control cells and in cells incubated for 36 h with 5 nM T3, 10 μM U0126, 10 μM SB203580, or 20 μM LY294002. (B) The Ras vector was cotransfected with the dominant-negative mutants pCEFL-N-rasasn17 (dnRas; 100 ng), pEF-myc-Raf301 (dnRaf; 500 ng), and pcDNA-dnMEK (dnMEK; 100 ng) or with the same amounts of the corresponding empty vectors. The luciferase activity was determined 36 h after transfection.

Journal:

Article Title: The Thyroid Hormone Receptor Is a Suppressor of ras -Mediated Transcription, Proliferation, and Transformation

doi: 10.1128/MCB.24.17.7514-7523.2004

Figure Lengend Snippet: The Ras/Erk pathway is the main effector for Ras-mediated stimulation of the cyclin D1 promoter in neuroblastoma cells. (A) N2a-β cells were transfected with the cyclin D1 promoter construct and 50 ng of Ha-rasval12. Reporter activity was determined in control cells and in cells incubated for 36 h with 5 nM T3, 10 μM U0126, 10 μM SB203580, or 20 μM LY294002. (B) The Ras vector was cotransfected with the dominant-negative mutants pCEFL-N-rasasn17 (dnRas; 100 ng), pEF-myc-Raf301 (dnRaf; 500 ng), and pcDNA-dnMEK (dnMEK; 100 ng) or with the same amounts of the corresponding empty vectors. The luciferase activity was determined 36 h after transfection.

Article Snippet: N2a-β cells were plated in 24-well plates and cotransfected by using Transfact transfection reagent (Promega) with 10 ng of a plasmid encoding enhanced green fluorescence protein (p-EGFP-C1; Clontech) and 40 ng of pCEFL-Ha- ras val12 or the corresponding empty vector.

Techniques: Transfection, Construct, Activity Assay, Incubation, Plasmid Preparation, Dominant Negative Mutation, Luciferase

CRE mediates stimulation by ras oncogene and repression by T3. (A) Schematic representation of the cyclin D1 promoter showing the position of binding sites for different transcription factors. In the lower panel, the effect of 36 h of incubation with 5 nM T3 on luciferase activity was determined in N2a-β cells cotransfected with Ha-rasval12 and the indicated 5′ cyclin D1 promoter deletions. (B) The CRE at nucleotide −58 in the promoter was mutated in the plasmid extending to −269, and the reporter activity was determined in the native (−269) and mutated (−269CREm) constructs after transfection with Ha-rasval12 and incubation in the presence or absence of T3. (C) CREB and ATF-2 bind the CRE of the cyclin D1 promoter. For the top panel, nuclear extracts from N2a-β cells were incubated in the absence (lane 2) or presence of antibodies for CREB, ATF-2, or a nonrelated protein (lanes 3, 4, and 5, respectively). Free probe (lane 1). For the bottom panel, EMSAs were performed with extracts from N2a-β cells transfected with 50 ng of Ha-rasval12 or the corresponding empty vector. Cells were incubated for 16 h in the presence or absence of 5 nM T3. (D) Cells were transfected with an UAS reporter plasmid and 250 ng of the indicated GAL4 fusion constructs or the GAL4 DBD alone. These constructs were cotransfected with Ha-rasval12 (50 ng) as indicated, and the luciferase activity was determined after 36 h in control and T3-treated cells.

Journal:

Article Title: The Thyroid Hormone Receptor Is a Suppressor of ras -Mediated Transcription, Proliferation, and Transformation

doi: 10.1128/MCB.24.17.7514-7523.2004

Figure Lengend Snippet: CRE mediates stimulation by ras oncogene and repression by T3. (A) Schematic representation of the cyclin D1 promoter showing the position of binding sites for different transcription factors. In the lower panel, the effect of 36 h of incubation with 5 nM T3 on luciferase activity was determined in N2a-β cells cotransfected with Ha-rasval12 and the indicated 5′ cyclin D1 promoter deletions. (B) The CRE at nucleotide −58 in the promoter was mutated in the plasmid extending to −269, and the reporter activity was determined in the native (−269) and mutated (−269CREm) constructs after transfection with Ha-rasval12 and incubation in the presence or absence of T3. (C) CREB and ATF-2 bind the CRE of the cyclin D1 promoter. For the top panel, nuclear extracts from N2a-β cells were incubated in the absence (lane 2) or presence of antibodies for CREB, ATF-2, or a nonrelated protein (lanes 3, 4, and 5, respectively). Free probe (lane 1). For the bottom panel, EMSAs were performed with extracts from N2a-β cells transfected with 50 ng of Ha-rasval12 or the corresponding empty vector. Cells were incubated for 16 h in the presence or absence of 5 nM T3. (D) Cells were transfected with an UAS reporter plasmid and 250 ng of the indicated GAL4 fusion constructs or the GAL4 DBD alone. These constructs were cotransfected with Ha-rasval12 (50 ng) as indicated, and the luciferase activity was determined after 36 h in control and T3-treated cells.

Article Snippet: N2a-β cells were plated in 24-well plates and cotransfected by using Transfact transfection reagent (Promega) with 10 ng of a plasmid encoding enhanced green fluorescence protein (p-EGFP-C1; Clontech) and 40 ng of pCEFL-Ha- ras val12 or the corresponding empty vector.

Techniques: Binding Assay, Incubation, Luciferase, Activity Assay, Plasmid Preparation, Construct, Transfection

TR represses Ha-rasval12-induced transformation of NIH 3T3 fibroblasts. (A) Representative foci formation in fibroblasts transfected with 50 ng of Ha-rasval12 and 4 μg of TRα1 or TRβ1. After transfection, cells were grown in the presence or absence of T3 for 14 days. In the right panel, the foci formed per dish were counted in three independent experiments. (B) Fibroblasts were transfected with 50 ng of Ha-rasval12 and 0.5 μg of TRβ1 in the presence or absence of 0.5 μg of a cyclin D1 expression vector. The cells were grown in hormone-depleted medium for the same time period, and the number of foci was scored in the different groups in control and T3-treated cultures.

Journal:

Article Title: The Thyroid Hormone Receptor Is a Suppressor of ras -Mediated Transcription, Proliferation, and Transformation

doi: 10.1128/MCB.24.17.7514-7523.2004

Figure Lengend Snippet: TR represses Ha-rasval12-induced transformation of NIH 3T3 fibroblasts. (A) Representative foci formation in fibroblasts transfected with 50 ng of Ha-rasval12 and 4 μg of TRα1 or TRβ1. After transfection, cells were grown in the presence or absence of T3 for 14 days. In the right panel, the foci formed per dish were counted in three independent experiments. (B) Fibroblasts were transfected with 50 ng of Ha-rasval12 and 0.5 μg of TRβ1 in the presence or absence of 0.5 μg of a cyclin D1 expression vector. The cells were grown in hormone-depleted medium for the same time period, and the number of foci was scored in the different groups in control and T3-treated cultures.

Article Snippet: N2a-β cells were plated in 24-well plates and cotransfected by using Transfact transfection reagent (Promega) with 10 ng of a plasmid encoding enhanced green fluorescence protein (p-EGFP-C1; Clontech) and 40 ng of pCEFL-Ha- ras val12 or the corresponding empty vector.

Techniques: Transformation Assay, Transfection, Expressing, Plasmid Preparation