Dataset Information

A refined, rapid and reproducible high resolution melt (HRM)-based method suitable for quantification of global LINE-1 repetitive element methylation.

ABSTRACT:

Background

The methylation of DNA is recognized as a key mechanism in the regulation of genomic stability and evidence for its role in the development of cancer is accumulating. LINE-1 methylation status represents a surrogate measure of genome-wide methylation.

Findings

Using high resolution melt (HRM) curve analysis technology, we have established an in-tube assay that is linear (r > 0.9986) with a high amplification efficiency (90-105%), capable of discriminating between partcipant samples with small differences in methylation, and suitable for quantifying a wide range of LINE-1 methylation levels (0-100%)--including the biologically relevant range of 50-90% expected in human DNA. We have optimized this procedure to perform using 2 μg of starting DNA and 2 ng of bisulfite-converted DNA for each PCR reaction. Intra- and inter-assay coefficients of variation were 1.44% and 0.49%, respectively, supporting the high reproducibility and precision of this approach.

Conclusions

In summary, this is a completely linear, quantitative HRM PCR method developed for the measurement of LINE-1 methylation. This cost-efficient, refined and reproducible assay can be performed using minimal amounts of starting DNA. These features make our assay suitable for high throughput analysis of multiple samples from large population-based studies.

SUBMITTER: Tse MY

PROVIDER: S-EPMC3284418 | biostudies-literature | 2011 Dec

REPOSITORIES: biostudies-literature

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Publications

A refined, rapid and reproducible high resolution melt (HRM)-based method suitable for quantification of global LINE-1 repetitive element methylation.

Tse M Yat MY Ashbury Janet E JE Zwingerman Nora N King Will D WD Taylor Sherry Am SA Pang Stephen C SC

BMC research notes 20111228

<h4>Background</h4>The methylation of DNA is recognized as a key mechanism in the regulation of genomic stability and evidence for its role in the development of cancer is accumulating. LINE-1 methylation status represents a surrogate measure of genome-wide methylation.<h4>Findings</h4>Using high resolution melt (HRM) curve analysis technology, we have established an in-tube assay that is linear (r > 0.9986) with a high amplification efficiency (90-105%), capable of discriminating between partci ...[more]

PMID: 22204640

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Dataset Information

A refined, rapid and reproducible high resolution melt (HRM)-based method suitable for quantification of global LINE-1 repetitive element methylation.

Background

Findings

Conclusions

Publications

A refined, rapid and reproducible high resolution melt (HRM)-based method suitable for quantification of global LINE-1 repetitive element methylation.

Similar Datasets

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