Genotyping
The process of ascertaining genetic variation data within individuals and populations, or genotyping, provides fundamental information regarding susceptibility, resistance, and progression of disease. Although there are different types of variants within the genome, the majority are single nucleotide polymorphisms (SNPs). Despite the fact that they account for <1% of the entire genome, SNPs represent nearly all differences between individuals, and have major implications toward disease predisposition. One of the largest genotyping efforts to date, the International HapMap Project, is currently capturing these inherited genetic variations through SNPs identification. Over three million of the estimated 10 million SNPs in the human genome have been validated thus far. Genotyping is a key enabling tool when investigating simple Mendelian disorders such as sickle-cell anemia, or understanding multigene interactions in complex diseases such as stroke, cancer, diabetes, and asthma. These techniques are heavily applied in human studies as well as in various animal models.
A vast array of assays and platforms exist today for genotype differentiation, and many laboratories use multiple approaches. End-point PCR techniques such as allele-specific amplification followed by gel analysis are useful when typing relatively small numbers of samples, and are often used as an entry point to molecular diagnostics. RT-PCR chemistries such as the popular TaqMan® assays are widely utilized for disease-association studies and population screens when numerous variants must be typed across a few thousand samples. As an alternative to RT-PCR, high-throughput Sanger sequencing is a frequently employed genotyping method as it offers a high level of selectivity and specificity. Microarrays are a common large-scale genotyping method employed in genome-wide association studies (GWAs). This approach allows researchers to interrogate hundreds of thousands of SNPs simultaneously, and has proven successful in identifying risk and protective factors related to Parkinson’s and Crohn’s disease as well as rheumatoid arthritis and mental illness. More recently, next-generation sequencing platforms are now also a viable genotyping option through sequence capture techniques, allowing the identification and sequence of multiple SNPs or other variant types in targeted regions of interest.
Caliper products enable genotyping laboratories to automate sample preparation and PCR reaction setup while also ensuring the quality of results. The Sciclone® G3 Workstation and Zephyr® Molecular Biology Workstation are ideal for DNA extraction, PCR reaction setup, and chip preparation for microarray experiments. In addition, Sciclone and Zephyr can be integrated with the Roche LightCycler and Twister II Plate Server for a complete, fully automated solution for RT-PCR applications. The LabChip® GX provides automated DNA sizing and quantitation for hundreds of samples in just a few hours. This platform is far superior to gel-based analysis of PCR products, and is essential in determining DNA quality prior to microarray and sequencing experiments. In addition, the LabChip GX can also be coupled with the Twister® II Plate Handler for longer walk away time and integrated into a Staccato® Workstation for complete automation of sample preparation and analysis.
LightCycler is a trademark of Roche Applied Science.
