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Rapid Identification of Promoter Hypermethylation in Hepatocellular Carcinoma by Pyrosequencing of Etiologically Homogeneous Sample Pools Emelyne Dejeux, Virginie Audard, Catherine Cavard, Ivo Glynne Gut, Benoit Terris, Jörg Tost The Journal of Molecular Diagnostics Volume 9, Issue 4, Pages (September 2007) DOI: /jmoldx Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions
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Figure 1 Pyrograms obtained by the analysis of mixtures with a known degree of methylation in the promoter of CDKN2A with 0% (A), 2% (B), 5% (C), and 10% (D) of methylation. E: The linearity of the signal for the sixth CpG position shown in A–D is demonstrated. Quantitative differences as low as 2 to 5% can be detected by pyrosequencing. Similar linear regression coefficients were obtained for the other five CpGs. CpG 1: y = 1.3 x + 2.3, R2 = 0.998; CpG 2: y = 1.57 x + 2.3, R2 = 0.997; CpG 3: y = 1.20 x + 1.0, R2 = 0.984; CpG 4: y = 0.88 x + 6.6, R2 = 0.978; CpG 5: y = 0.79 x + 4.2, R2 = F: The linearity of the signal throughout the entire dynamic range (0 to 100%) for the sixth CpG position is shown. All assays were performed in triplicate. The resolution and linear correlation is similar for IGF2 as demonstrated in G. The Journal of Molecular Diagnostics 2007 9, DOI: ( /jmoldx ) Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions
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Figure 2 Graphical representation (boxplots) of the results obtained by the analysis of the individual samples for 112 CpGs in five genes (CDKN2A, GSTP1, MLH1, IGF2, and CTNNB1) and the level of statistical significance for differential methylation values. *P < 0.05, **P < 0.005, and ***P < The Journal of Molecular Diagnostics 2007 9, DOI: ( /jmoldx ) Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions
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