On lab techniques for studying genetic variation, a review


Our emphasis at OpenHelix is to provide training on how to use specific genomics online tools and databases that you need for your research. We are sometimes asked, though, generic cialis 10mg for some more theoretical aspects of the research such the understanding the methodologies of SNP analysis or sequence alignment. So to help answer these questions here on the OpenHelix blog, we will be offering occasional links and reviews of videos and other instructional material on the web for these more theoretical aspects of genomics research. The OpenHelix Blog is pleased to welcome Cyndy Perrealt-Micale as a regular contributor for this feature.

Karen Moleke Talk

For those of you interested in the latest developments in laboratory techniques available to study genetic variability, there is a great lecture available by Karen Mohlke of the University of North Carolina. No background knowledge is needed to understand the topics discussed here.

Dr. Mohlke effectively reviews and explains all the major types of genetic variations, and the theory behind the latest methods used to study them.

Even those of us that are a bit rusty in this field can gain a great deal of knowledge from this because of the way she leads you from a discussion of the basic background right to in depth descriptions of the contemporary laboratory techniques used to determine and quantify many types of polymorphisms. Human SNPs are highlighted the most in Karin’s talk, and this certainly makes the target audience for this lecture quite broad. Anyone interested in the discovery of genes that may have medically significant consequences could benefit from listening to this. Advantages, disadvantages and the rationale behind the methodology are clearly and concisely presented, and their applicability and cost-effectiveness for large scale and small scale genotype analysis are presented. If you want to learn about the techniques used currently for determining genetic variability (whether it is for a single experimental determination or for tens of thousands of samples) then this is one lecture you may not want to miss!

The major classes of genetic variations, including microsatellites, SNPs, indels (insertion/deletion polymorphs) and structural variations are presented with specific examples, overviews and frequencies of occurrence. She reviews the criteria one can use for method selection such as cost-effectiveness, multiplex ability, assay development time and the ability to automate steps. An overview of available SNP genotyping methods, including the theoretical background, and its advantages and disadvantages in terms of sample size, speed, reliability and many other variables are discussed. This I found to be a very useful and enlightening discussion. For example, one can learn that currently the easiest method for smaller samples sizes is sequencing, yet the disadvantages are that this can be very expensive for larger sample numbers and some of the data acquired may be ambiguous and require multiple reads. Hybridization, mini-sequencing, including both pyrosequencing and primer extension mass spectrometry, allele-specific PCR and oligonucleotide ligation assays are all presented and compared to each other with similar detail.

Quality control of genotyping data is an important concern that is also addressed here. Some of the methods discussed to improve the quality of data are the inclusion of duplicate samples, comparing your results to those already in databases (like HapMap which includes data for 4 million SNPs currently), negative controls, analysis by a different experimental platform and using the Hardy-Weinberg equation to compare your observed genotype frequency to the expected frequency.

Several large scale genotyping techniques are discussed, including Affymetrix GeneChip arrays and Illumina Infinium assays. These are generally geared towards the analysis of 10,000 to 1 million SNPs at the same time and, similarly to the smaller scale methods, all of their disadvantages and advantages are presented with appropriate detail. Statistics are shown regarding the percent of genome coverage by these various large scale techniques, and Dr. Mohlke informs us that simply looking at the percentage of coverage is not enough. There are differences in local coverage that we should be aware of because this results in certain areas of the genome being underrepresented. An interesting new development is that some of these large scale techniques designed for SNP analysis are also able to detect variations in copy numbers. This is one area that may be focused on in the future as more probes are being added to these chips to help detect both copy number changes and SNPs in one experiment.

In Karen’s opinion, the future direction of laboratory technique development for genotyping will be heading towards faster, easier and cheaper methods, such as the more predominant use of next generation sequencing for both rare and common SNPs. These technological advancements should lead to improved maps for structural variations and the discovery of many new susceptibility genes for complex traits.