Microsatellite Instability
Microsatellite instability describes the reduced fidelity during the replication of repetitive DNA often occurring in tumor cells. It's thought to be caused by strand slippage during DNA replication due to mutations in DNA mismatch repair genes. MSI leads to the appearance of multiple alleles at microsatellite loci, which can be easily identified. Mutations in microsatellite regions are termed MSI, while replication error (RER) is usually defined as MSI at multiple microsatellite markers.
The electropherogram of the dinucleotide repeat marker D18S35 from a homozygous individual (Figure 1). The appearance of numerous extra alleles at lower molecular weights in the tumor sample (Figure 1, bottom panel) indicates significant genomic instability.
Experimental Workflow: Find the Products You Need for Every Step
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Step 1: DNA Purification
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.
Products for Step 1: DNA Purification
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DNA Isolation Kits
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Step 2: Primer Design & PCR Amplification
Amplify microsatellite loci of interest with fluorescently labeled primer pair.
Products for Step 2: Primer Design & PCR Amplification
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Custom Microsatellite Primers
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Step 3: Sample Prep for Fragment Analysis Run
Add aliquot of PCR reaction to size standard and Hi-Di Formamide mix; heat denature and proceed with electrophoresis.
Products for Step 3: Sample Prep for Fragment Analysis Run
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Hi-Di™ Formamide
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Highly deionized formamide used to resuspend samples before electrokinetic injection in capillary electrophoresis systems.
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GeneScan™ Size Standards
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GeneScan™ Size Standards are fluorescently-labeled DNA fragments required for performing fragment analysis and genotyping applications on our capillary electrophoresis platforms.
Step 4: Capillary Electrophoresis
During capillary electrophoresis, the products of the PCR reaction are injected electrokinetically into capillaries filled with polymer. High voltage is applied so that the negatively charged DNA fragments move through the polymer in the capillaries toward the positive electrode.
Products for Step 4: Capillary Electrophoresis
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3500 Genetic Analyzer
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The 3500 8-capillary platform can run a wide variety of applications—including de novo sequencing and resequencing (mutational profiling)—as well as microsatellite analysis, MLPA™, LOH, MLST, AFLP® and SNP validation or screening. The majority of applications can be run on a single polymer and capillary array.
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3500xL Genetic Analyzer
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The 3500xL 24-capillary platform can run a wide variety of applications, including de novo sequencing and resequencing (mutational profiling), as well as microsatellite analysis, MLPA™, LOH, MLST, AFLP® and SNP validation or screening. The majority of applications can be run on a single polymer and capillary array.
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3730 DNA Analyzer
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The 48-capillary 3730 DNA Analyzer is the Gold Standard in medium-to-high throughput genetic analysis. Use this for DNA fragment analysis applications such as microsatellites, AFLP, SNP analysis, mutation detection and traditional DNA sequencing. Get the highest quality data at a low cost per sample. This is upgradeable to 96 capillaries.
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3730xl DNA Analyzer
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The 96-capillary 3730xl DNA Analyzer is the Gold Standard for high throughput genetic analysis. Use this for DNA fragment analysis applications such as microsatellites, AFLP, SNP analysis, mutation detection and traditional DNA sequencing. Get the highest quality data at a low cost per sample.
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3130 Genetic Analyzer
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Applied Biosystems 3130 Genetic Analyzer is the latest generation of 4-capillary electrophoresis instruments for the low to medium throughput laboratories. The system offers industry-leading performance, plus sophisticated automation capabilities allowing you to save time, reduce costs and increase productivity.
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3130xl Genetic Analyzer
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Applied Biosystems 3130xl Genetic Analyzer is the latest generation of 16-capillary electrophoresis instruments for the medium throughput laboratories.
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310 Genetic Analyzer
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The ABI PRISM® 310 Genetic Analyzer is an automated single-capillary genetic analyzer designed for a wide range of sequencing and fragment analysis applications.
Step 5: Data Analysis
Analyze microsatellite loci for multiple alleles and the appearance of numerous extra alleles indicative of genomic instability.
Products for Step 5: Data Analysis
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GeneMapper® Software
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GeneMapper is a flexible genotyping software package that provides DNA sizing and quality allele calls for all Applied Biosystems electrophoresis-based genotyping systems.
