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Figure 1: Cumulative Estimation for Methylation State of CpG Islands within an Amplicon. Methylated fragments migrate faster (Panel A, B) than those that are unmethylated (Panel C).

MSMSA is a DNA fragment analysis application used to assess the degree of methylation present in a given amplicon.

Samples from a fragment analysis assay can also be used in a downstream sequencing step, making this application ideal for screening. The assay primers are designed in regions that don’t contain CpGs using Methyl Primer Express® Software, so that PCR amplification is not dependent on methylation status.

Step-by-Step Guide to Methylation Sensitive Mobility Shift Assay

DNA Purification kit

DNA extraction is a critical first step in the experimental workflow of DNA Sequencing and Fragment analysis. The overall quality, accuracy and length of the DNA sequence read can be significantly affected by characteristics of the sample itself, and the method chosen for nucleic acid extraction. Ideal methods will vary depending on the source or tissue type, how it was obtained from its source, and how the sample was handled or stored prior to extraction.

Recommended Products: DNA Isolation

PCR Kit

The methylSEQr™ Bisulfite Conversion Kit converts non-methylated cytosines (C) in the DNA sample to uracil (U). Methylated cytosines remain unchanged. Comparing the sequences of treated vs. non-treated DNA and detection of converted cytosines (now apparent as T base) determine which cytosines in the sample are methylated in CpG dinucleotides.

Recommended Products: Perform Bisulfate Conversion

Which Kits are Right for You?

PCR Kit

Design high quality PCR primers for methylation mapping experiments. Simply cut and paste in your region of interest in the primer design software. The tool searches for CpG islands and simulates bisulfite modification of DNA in silico. Perform PCR to amplify region of interest.

Recommended Products: Perform PCR

Learn MoreRecommended Products
Fragment Analysis sizing graph image.

Add aliquot of PCR product to size standard and Hi-Di Formamide mix; heat denature and proceed with electrophoresis.

Recommended Products: Prepare Sample for Analysis

Sizing Information

To learn more about our GeneScan Size standards used in sizing experiments please see our table below.

When DNA fragments are labeled with: Choose a size standards labeled with: Recommended Size Standard Possible Applications Other Kits and Products
dR110, dR6G, dTAMRA™, dROX (Dye Set DS-02 - Filter E5) LIZ®
GeneScan™ 120 LIZ® Size Standard
GeneScan™ 600 LIZ Size Standard v2.0
SNaPshot®
Microsatellite, LOH, ISSR, Chimerism, ISSR, RFLP, T-RFLP, MSMSA
SNaPshot® primer focus kits
Custom Labeled Primer
5-FAM™, HEX™, NED™ (Dye Set DS-30 - Filter D) ROX™ GeneScan™ 500 ROX™ Size Standard Custom Fragment Analysis Custom Labeled Primer
6-FAM™, VIC®, NED™ (Dye Set DS-31 - Filter D**) ROX™ GeneScan™ 500 ROX™ Size Standard

Custom Fragment Analysis

Custom Labeled Primer
5-FAM™, JOE, NED™ (Dye Set DS-32 - Filter F) ROX™ GeneScan™ 500 ROX™ Size Standard
Microsatellite (Forensic)
AFLP
AmpFℓSTR® Kits,
Plant/Microbial AFLP Kits®
6-FAM™, VIC®, NED™, PET® (Dye Set DS-33 - Filter G5) LIZ® GeneScan™ 600 LIZ Size Standard v2.0
Microsatellite (Forensic)
Custom Fragment Analysis
AmpFℓSTR® Kits,
Custom Labeled Primer
Capillary Electrophoresis

During capillary electrophoresis, the products of the PCR are injected electrokinetically into capillaries filled with polymer. High voltage is applied so that the fluorescent DNA fragments are separated by size and are detected by a laser/camera system.

Which Electrophoresis Instrument (Genetic Analyzer) Is Right for You?

  Instrument
 3730xl
3730xL
3730
3730
3500xl
3500xL
3500
3500
3130xl
3130xL
3130
3130
310
310
Number of Capillaries96482481641
Compatible Applications:
(S) Supported; (A) AB Demonstrated; (C) Customer Demonstrated; (N) Not Supported
MethylationSSSSSSC

Analyze the MSMSA profile.

Which Data Analysis Software Is Right for You?

Highlighted products for Methylation Sensitive Mobility Shift Assay:

Verti Thermal Cycler

3500 Series Genetic Analyzers

GeneMapper® Software