Methylation-sensitive high-resolution melting (MS-HRM) is a PCR-based in-tube method for detecting methylation levels at specific loci of interest. The unique primer design facilitates the high sensitivity of the assay, enabling detection of methylation alleles as low as 0.1-1% in an unmethylated background. Creative BioMart offers an MS-HRM solution, which is based on DNA high-resolution melting technology for high sensitivity, labor- and cost-effective single-locus methylation studies.

What Is MS-HRM?

The principle of detecting DNA methylation at a specific locus of interest using MS-HRM is based on specific primers design and measurement of differences in melting characteristics between methylated and unmethylated alleles after bisulfite conversion. Briefly, after the bisulfite conversion, the DNA is subjected to PCR in which a saturating fluorescent dye is present. When bound to double-stranded DNA, the dye will omit fluorescence, allowing the amplification process to be tracked during PCR. After a certain number of cycles, the temperature is raised to dissociate the amplicons. As the temperature gradually increases, the double helix dissociates and releases the embedded dye, thereby promoting the reduction of fluorescence. Amplicon from a template with high levels of methylation, containing a high proportion of cytosine (C) and guanine (G) after bisulfite conversion, will melt at a higher temperature than templates from unmethylated DNA with high adenine (A) and thymine (T) content. Therefore, the quantitative result of DNA methylation can be known by real-time analysis of the fluorescent signal.

The schematic illustration of the principle behind MS-HRM is as follows. Primers designed for MS-HRM are complementary to the methylated template, but a specific annealing temperature ensures binding to both the methylated and the unmethylated templates. Only the CpG sites between the primers are assessed for their methylation status. A schematic illustration of the resulting melting curve is shown for a fully methylated template (red curve), an unmethylated template (blue), and a heterogeneously methylated template (green).

Figure 1. Schematic illustration of the principle behind MS-HRM. (Dianna Hussmann.; Lise Lotte Hansen. 2018). Primers (blue arrows); filled black circles (methylated cytosines), filled gray circles (methylated or unmethylated cytosines), open circles (unmethylated cytosines).

Features of MS-HRM

1. Highly sensitive and capable of detecting methylation status of low copy number DNA from highly degraded materials
2. Carried out in a closed tube, limiting the risk of contamination
3. Suitable for noninvasive detection of potential methylation-specific biomarkers
4. High throughput
5. Cost-effective technique

Our Advantages

1. Tolerance of multiple sample types: we accept a wide range of biological and clinical samples including genomic DNA, cells, tissues, formalin-fixed and paraffin-embedded samples. Sample sources include human, animals, plants and microorganisms.
2. One-stop service: customers only need to provide samples, and we will complete full MS-HRM experiment from specific primer design, standard curve construction, bisulfite conversion, on-machine detection to data analysis. The complete report is provided to you with both raw and analyzed data, graphs, detailed protocols, and a summary of findings.
3. Skillful technique team and first-class technology platform: the specific primers we designed to facilitate the amplification of methylated alleles will confer high sensitivity to MS-HRM assays, and the ability to detect methylation levels between 0.1% and 1%, in addition, using high concentrations of saturated fluorescent dyes to guarantee high resolution. Our fully automatic fluorescent quantitative PCR system can precisely control temperature changes and has advanced data acquisition function.
4. Professional data analysis: with a professional data analysis platform designed for HRM, Creative BioMart provides you with a full range of data analysis services.

Workflow of MS-HRM Analysis Service at Creative BioMart

Figure 2. Workflow of MS-HRM analysis service at Creative BioMart

As a supplier focused on epigenetic research services, Creative BioMart has more than 10 years of experience in DNA methylation analysis. With our extensive expertise and state-of-the-art technology platform, we will provide our customers with high quality services at the most competitive price. Our services are tailored to meet your specific needs. contact us for more information about our MS-HRM analysis service.

References
1. Wojdacz T K.; et al. Methylation-sensitive high-resolution melting. Nature Protocols. 2008, 3(12): 1903-1908.
2. Dianna Hussmann.; Lise Lotte Hansen. Methylation-sensitive high resolution melting (MS-HRM). DNA Methylation Protocols. 2018.