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pgl2 promoter control vector  (Promega)

 
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    Promega pgl2 promoter control vector
    A , Expression of CDO1 in CRC cell lines was examined by RT-PCR analysis. No basal expression of CDO1 was seen in all CRC cell lines, and all these cell lines harbored CDO1 methylation (M). Silenced CDO1 was reactivated after treatment with the demethylating agent, 5-Aza-dC, indicating that CDO1 methylation correlates tightly with loss of gene expression in CRC cell lines. β-actin was used as a loading control. m, cells treated with vehicle only; a, cells treated with 5-Aza-dC (5 µM for three days). L, 1 Kb Plus DNA ladder. B , Methylation status of individual CpGs of the island in 5 CRC cell lines and 21 pairs of primary CRC (PT) and their corresponding normal colon tissues (PN) is shown. A total of 38 CGs were numbered from the first to last CG in the sequences as indicated. Black circle, methylation; white circle, unmethylation; grey circle, co-existence of methylated and unmethylated alleles; dashed circle, undetermined. C , Analysis of CDO1 promoter activity by luciferase reporter assay in CDO1 -negative (HCT116) and -positive (HEK293) cells. The promoter constructs <t>(pGL2-</t> CDO1 -#1 and -#2) were pre-treated with or without Sss I methylase for 8 hrs before transfection. High activity of the CDO1 promoter was detected in HEK293 where CDO1 was expressed. Data are expressed as fold increase over pGL2-basic activity. Experiments were done in triplicate, and values indicate means ± SD. Mean values are presented. D. Scatter plot of CDO1 methylation levels in tissues and cell lines (CL) (left). TaqMan methylation values (TaqMeth V) is described in Materials and Method. TaqMan-MSP was performed in duplicate format, and experiments were repeated twice. Data showed reproducible and concordant results. PT, primary CRC; PN, matched normal colon tissues from colon cancer patients; NN, normal colon epithelium from non-cancer patients. Line indicates the optimal cut-off value for CDO1 calculated from ROC analysis. Sample numbers showing TaqMeth V over the cut-off are indicated. The overall TaqMeth V detected in PT was significantly higher than that in PN (right). TaqMan value of two NN (22%, 2/9) was above the cut-off value (24.56 and 17.86 each), suggesting that a low level of CDO1 methylation can be caused by other unknown mechanisms. E , ROC curve analysis of TaqMeth V of CDO1 in CRC. The Area under the ROC (AUROC) conveys the accuracy in distinguishing matched normal colon (PN) from CRC (PT) in terms of its sensitivity and specificity ( P<0.001 ). Solid line, CDO1 ; dashed line, no discrimination. F , Methylation levels of normal (PN) and tumor tissues (PT) in individual patients.
    Pgl2 Promoter Control Vector, 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/pgl2 promoter control vector/product/Promega
    Average 90 stars, based on 1 article reviews
    pgl2 promoter control vector - by Bioz Stars, 2026-05
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    Images

    1) Product Images from "Cysteine Dioxygenase 1 Is a Tumor Suppressor Gene Silenced by Promoter Methylation in Multiple Human Cancers"

    Article Title: Cysteine Dioxygenase 1 Is a Tumor Suppressor Gene Silenced by Promoter Methylation in Multiple Human Cancers

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0044951

    A , Expression of CDO1 in CRC cell lines was examined by RT-PCR analysis. No basal expression of CDO1 was seen in all CRC cell lines, and all these cell lines harbored CDO1 methylation (M). Silenced CDO1 was reactivated after treatment with the demethylating agent, 5-Aza-dC, indicating that CDO1 methylation correlates tightly with loss of gene expression in CRC cell lines. β-actin was used as a loading control. m, cells treated with vehicle only; a, cells treated with 5-Aza-dC (5 µM for three days). L, 1 Kb Plus DNA ladder. B , Methylation status of individual CpGs of the island in 5 CRC cell lines and 21 pairs of primary CRC (PT) and their corresponding normal colon tissues (PN) is shown. A total of 38 CGs were numbered from the first to last CG in the sequences as indicated. Black circle, methylation; white circle, unmethylation; grey circle, co-existence of methylated and unmethylated alleles; dashed circle, undetermined. C , Analysis of CDO1 promoter activity by luciferase reporter assay in CDO1 -negative (HCT116) and -positive (HEK293) cells. The promoter constructs (pGL2- CDO1 -#1 and -#2) were pre-treated with or without Sss I methylase for 8 hrs before transfection. High activity of the CDO1 promoter was detected in HEK293 where CDO1 was expressed. Data are expressed as fold increase over pGL2-basic activity. Experiments were done in triplicate, and values indicate means ± SD. Mean values are presented. D. Scatter plot of CDO1 methylation levels in tissues and cell lines (CL) (left). TaqMan methylation values (TaqMeth V) is described in Materials and Method. TaqMan-MSP was performed in duplicate format, and experiments were repeated twice. Data showed reproducible and concordant results. PT, primary CRC; PN, matched normal colon tissues from colon cancer patients; NN, normal colon epithelium from non-cancer patients. Line indicates the optimal cut-off value for CDO1 calculated from ROC analysis. Sample numbers showing TaqMeth V over the cut-off are indicated. The overall TaqMeth V detected in PT was significantly higher than that in PN (right). TaqMan value of two NN (22%, 2/9) was above the cut-off value (24.56 and 17.86 each), suggesting that a low level of CDO1 methylation can be caused by other unknown mechanisms. E , ROC curve analysis of TaqMeth V of CDO1 in CRC. The Area under the ROC (AUROC) conveys the accuracy in distinguishing matched normal colon (PN) from CRC (PT) in terms of its sensitivity and specificity ( P<0.001 ). Solid line, CDO1 ; dashed line, no discrimination. F , Methylation levels of normal (PN) and tumor tissues (PT) in individual patients.
    Figure Legend Snippet: A , Expression of CDO1 in CRC cell lines was examined by RT-PCR analysis. No basal expression of CDO1 was seen in all CRC cell lines, and all these cell lines harbored CDO1 methylation (M). Silenced CDO1 was reactivated after treatment with the demethylating agent, 5-Aza-dC, indicating that CDO1 methylation correlates tightly with loss of gene expression in CRC cell lines. β-actin was used as a loading control. m, cells treated with vehicle only; a, cells treated with 5-Aza-dC (5 µM for three days). L, 1 Kb Plus DNA ladder. B , Methylation status of individual CpGs of the island in 5 CRC cell lines and 21 pairs of primary CRC (PT) and their corresponding normal colon tissues (PN) is shown. A total of 38 CGs were numbered from the first to last CG in the sequences as indicated. Black circle, methylation; white circle, unmethylation; grey circle, co-existence of methylated and unmethylated alleles; dashed circle, undetermined. C , Analysis of CDO1 promoter activity by luciferase reporter assay in CDO1 -negative (HCT116) and -positive (HEK293) cells. The promoter constructs (pGL2- CDO1 -#1 and -#2) were pre-treated with or without Sss I methylase for 8 hrs before transfection. High activity of the CDO1 promoter was detected in HEK293 where CDO1 was expressed. Data are expressed as fold increase over pGL2-basic activity. Experiments were done in triplicate, and values indicate means ± SD. Mean values are presented. D. Scatter plot of CDO1 methylation levels in tissues and cell lines (CL) (left). TaqMan methylation values (TaqMeth V) is described in Materials and Method. TaqMan-MSP was performed in duplicate format, and experiments were repeated twice. Data showed reproducible and concordant results. PT, primary CRC; PN, matched normal colon tissues from colon cancer patients; NN, normal colon epithelium from non-cancer patients. Line indicates the optimal cut-off value for CDO1 calculated from ROC analysis. Sample numbers showing TaqMeth V over the cut-off are indicated. The overall TaqMeth V detected in PT was significantly higher than that in PN (right). TaqMan value of two NN (22%, 2/9) was above the cut-off value (24.56 and 17.86 each), suggesting that a low level of CDO1 methylation can be caused by other unknown mechanisms. E , ROC curve analysis of TaqMeth V of CDO1 in CRC. The Area under the ROC (AUROC) conveys the accuracy in distinguishing matched normal colon (PN) from CRC (PT) in terms of its sensitivity and specificity ( P<0.001 ). Solid line, CDO1 ; dashed line, no discrimination. F , Methylation levels of normal (PN) and tumor tissues (PT) in individual patients.

    Techniques Used: Expressing, Reverse Transcription Polymerase Chain Reaction, Methylation, Activity Assay, Luciferase, Reporter Assay, Construct, Transfection



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    A , Expression of CDO1 in CRC cell lines was examined by RT-PCR analysis. No basal expression of CDO1 was seen in all CRC cell lines, and all these cell lines harbored CDO1 methylation (M). Silenced CDO1 was reactivated after treatment with the demethylating agent, 5-Aza-dC, indicating that CDO1 methylation correlates tightly with loss of gene expression in CRC cell lines. β-actin was used as a loading control. m, cells treated with vehicle only; a, cells treated with 5-Aza-dC (5 µM for three days). L, 1 Kb Plus DNA ladder. B , Methylation status of individual CpGs of the island in 5 CRC cell lines and 21 pairs of primary CRC (PT) and their corresponding normal colon tissues (PN) is shown. A total of 38 CGs were numbered from the first to last CG in the sequences as indicated. Black circle, methylation; white circle, unmethylation; grey circle, co-existence of methylated and unmethylated alleles; dashed circle, undetermined. C , Analysis of CDO1 promoter activity by luciferase reporter assay in CDO1 -negative (HCT116) and -positive (HEK293) cells. The promoter constructs <t>(pGL2-</t> CDO1 -#1 and -#2) were pre-treated with or without Sss I methylase for 8 hrs before transfection. High activity of the CDO1 promoter was detected in HEK293 where CDO1 was expressed. Data are expressed as fold increase over pGL2-basic activity. Experiments were done in triplicate, and values indicate means ± SD. Mean values are presented. D. Scatter plot of CDO1 methylation levels in tissues and cell lines (CL) (left). TaqMan methylation values (TaqMeth V) is described in Materials and Method. TaqMan-MSP was performed in duplicate format, and experiments were repeated twice. Data showed reproducible and concordant results. PT, primary CRC; PN, matched normal colon tissues from colon cancer patients; NN, normal colon epithelium from non-cancer patients. Line indicates the optimal cut-off value for CDO1 calculated from ROC analysis. Sample numbers showing TaqMeth V over the cut-off are indicated. The overall TaqMeth V detected in PT was significantly higher than that in PN (right). TaqMan value of two NN (22%, 2/9) was above the cut-off value (24.56 and 17.86 each), suggesting that a low level of CDO1 methylation can be caused by other unknown mechanisms. E , ROC curve analysis of TaqMeth V of CDO1 in CRC. The Area under the ROC (AUROC) conveys the accuracy in distinguishing matched normal colon (PN) from CRC (PT) in terms of its sensitivity and specificity ( P<0.001 ). Solid line, CDO1 ; dashed line, no discrimination. F , Methylation levels of normal (PN) and tumor tissues (PT) in individual patients.
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    Image Search Results


    A , Expression of CDO1 in CRC cell lines was examined by RT-PCR analysis. No basal expression of CDO1 was seen in all CRC cell lines, and all these cell lines harbored CDO1 methylation (M). Silenced CDO1 was reactivated after treatment with the demethylating agent, 5-Aza-dC, indicating that CDO1 methylation correlates tightly with loss of gene expression in CRC cell lines. β-actin was used as a loading control. m, cells treated with vehicle only; a, cells treated with 5-Aza-dC (5 µM for three days). L, 1 Kb Plus DNA ladder. B , Methylation status of individual CpGs of the island in 5 CRC cell lines and 21 pairs of primary CRC (PT) and their corresponding normal colon tissues (PN) is shown. A total of 38 CGs were numbered from the first to last CG in the sequences as indicated. Black circle, methylation; white circle, unmethylation; grey circle, co-existence of methylated and unmethylated alleles; dashed circle, undetermined. C , Analysis of CDO1 promoter activity by luciferase reporter assay in CDO1 -negative (HCT116) and -positive (HEK293) cells. The promoter constructs (pGL2- CDO1 -#1 and -#2) were pre-treated with or without Sss I methylase for 8 hrs before transfection. High activity of the CDO1 promoter was detected in HEK293 where CDO1 was expressed. Data are expressed as fold increase over pGL2-basic activity. Experiments were done in triplicate, and values indicate means ± SD. Mean values are presented. D. Scatter plot of CDO1 methylation levels in tissues and cell lines (CL) (left). TaqMan methylation values (TaqMeth V) is described in Materials and Method. TaqMan-MSP was performed in duplicate format, and experiments were repeated twice. Data showed reproducible and concordant results. PT, primary CRC; PN, matched normal colon tissues from colon cancer patients; NN, normal colon epithelium from non-cancer patients. Line indicates the optimal cut-off value for CDO1 calculated from ROC analysis. Sample numbers showing TaqMeth V over the cut-off are indicated. The overall TaqMeth V detected in PT was significantly higher than that in PN (right). TaqMan value of two NN (22%, 2/9) was above the cut-off value (24.56 and 17.86 each), suggesting that a low level of CDO1 methylation can be caused by other unknown mechanisms. E , ROC curve analysis of TaqMeth V of CDO1 in CRC. The Area under the ROC (AUROC) conveys the accuracy in distinguishing matched normal colon (PN) from CRC (PT) in terms of its sensitivity and specificity ( P<0.001 ). Solid line, CDO1 ; dashed line, no discrimination. F , Methylation levels of normal (PN) and tumor tissues (PT) in individual patients.

    Journal: PLoS ONE

    Article Title: Cysteine Dioxygenase 1 Is a Tumor Suppressor Gene Silenced by Promoter Methylation in Multiple Human Cancers

    doi: 10.1371/journal.pone.0044951

    Figure Lengend Snippet: A , Expression of CDO1 in CRC cell lines was examined by RT-PCR analysis. No basal expression of CDO1 was seen in all CRC cell lines, and all these cell lines harbored CDO1 methylation (M). Silenced CDO1 was reactivated after treatment with the demethylating agent, 5-Aza-dC, indicating that CDO1 methylation correlates tightly with loss of gene expression in CRC cell lines. β-actin was used as a loading control. m, cells treated with vehicle only; a, cells treated with 5-Aza-dC (5 µM for three days). L, 1 Kb Plus DNA ladder. B , Methylation status of individual CpGs of the island in 5 CRC cell lines and 21 pairs of primary CRC (PT) and their corresponding normal colon tissues (PN) is shown. A total of 38 CGs were numbered from the first to last CG in the sequences as indicated. Black circle, methylation; white circle, unmethylation; grey circle, co-existence of methylated and unmethylated alleles; dashed circle, undetermined. C , Analysis of CDO1 promoter activity by luciferase reporter assay in CDO1 -negative (HCT116) and -positive (HEK293) cells. The promoter constructs (pGL2- CDO1 -#1 and -#2) were pre-treated with or without Sss I methylase for 8 hrs before transfection. High activity of the CDO1 promoter was detected in HEK293 where CDO1 was expressed. Data are expressed as fold increase over pGL2-basic activity. Experiments were done in triplicate, and values indicate means ± SD. Mean values are presented. D. Scatter plot of CDO1 methylation levels in tissues and cell lines (CL) (left). TaqMan methylation values (TaqMeth V) is described in Materials and Method. TaqMan-MSP was performed in duplicate format, and experiments were repeated twice. Data showed reproducible and concordant results. PT, primary CRC; PN, matched normal colon tissues from colon cancer patients; NN, normal colon epithelium from non-cancer patients. Line indicates the optimal cut-off value for CDO1 calculated from ROC analysis. Sample numbers showing TaqMeth V over the cut-off are indicated. The overall TaqMeth V detected in PT was significantly higher than that in PN (right). TaqMan value of two NN (22%, 2/9) was above the cut-off value (24.56 and 17.86 each), suggesting that a low level of CDO1 methylation can be caused by other unknown mechanisms. E , ROC curve analysis of TaqMeth V of CDO1 in CRC. The Area under the ROC (AUROC) conveys the accuracy in distinguishing matched normal colon (PN) from CRC (PT) in terms of its sensitivity and specificity ( P<0.001 ). Solid line, CDO1 ; dashed line, no discrimination. F , Methylation levels of normal (PN) and tumor tissues (PT) in individual patients.

    Article Snippet: The pGL2 promoter control vector (Promega, Madison, WI) was digested with both KpnI and XhoI and treated with calf intestinal alkaline phosphatase.

    Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Methylation, Activity Assay, Luciferase, Reporter Assay, Construct, Transfection