Open Access Open Badges Technical Note

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

M Yat Tse1*, Janet E Ashbury2, Nora Zwingerman2, Will D King2, Sherry AM Taylor3 and Stephen C Pang1*

Author Affiliations

1 Department of Anatomy and Cell Biology, Queen's University, Kingston, ON, Canada

2 Department of Community Health and Epidemiology, Queen's University, Kingston, ON, Canada

3 Department of Laboratory Medicine, Saint John Regional Hospital, Horizon Health Network and Department of Pathology, Dalhousie University, Halifax, NS, Canada

For all author emails, please log on.

BMC Research Notes 2011, 4:565  doi:10.1186/1756-0500-4-565

Published: 28 December 2011



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.


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.


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.