Exome sequencing (or whole-exome sequencing/WES) is a term that identifies the sequencing of the entire coding region of the genome of an individual. Until a few years ago, genetic testing for excellence consisted of the sequencing of a single gene or a small group of genes using the method of capillary electrophoresis, more commonly known as Sanger sequencing. The advent of new sequencing technologies (Next Generation Sequencing - NGS and Third Generation Sequencing) has instead turned to the study of the genetics of human disease, leading to an era of unprecedented productivity. Due to the costs and reduced times, via NGS it is now possible to analyze a large number of DNA fragments in parallel to obtain the sequence of many genes simultaneously or even the entire coding region of an individual. This is of great interest for both routine diagnostics and scientific research. In fact, even if the coding region represents only 1% of the entire genome, it is estimated that up to 85% of all pathogenic mutations are contained in this region. Exome sequencing is useful for example in those cases where a a precise clinical diagnosis is lacking or in genetically heterogeneous disorders, for which exome sequencing can help identifying yet unknown causative mutations.
As said, by definition, exome sequencing refers to the sequencing of the coding region of the genome. It is true that some enrichment kits are still incomplete and lack coverage of some areas, however it is also true that other kits containing primers for some non-coding regions, such as the exon/intron boundaries (where lots of mutations can be found), are already available.
EXOME SEQUENCING APPLICATIONS
1. STUDY OF MENDELIAN DISORDERS
The traditional approach in Mendelian pathology research has always been the linkage study followed by positional cloning. This approach, however, presents considerable difficulties, mainly related to the need of having to test more than one family (not an easy undertaking for very rare diseases) and genetic heterogeneity that sometimes characterizes certain Mendelian diseases. Think, for example, of syndromes such as Bardet-Biedl, Joubert or Kallmann, which can be caused by mutation in one of several different genes. Exome sequencing, however, screening thoroughly the coding sequence, allows you to quickly identify pathogenic or likely pathogenic variants also in genetically heterogeneous disorders. The genes involved in Kabuki and Miller syndromes pathogenesis, for example, have recently been identified thanks to exome sequencing studies. Suffice it to say that, only in the last three years, the genes associated to about 100 Mendelian diseases have been discovered through exome sequencing!
2. MULTIFACTORIAL DISEASES STUDIES
In recent years, the traditional approach to the genetic study of multifactorial diseases (i.e. those diseases that are supposed to arise as a result of the interaction between environmental factors and genetic predisposition) was mainly based on genome array, i.e. screening of a selected frequent variants scattered throughout the genome. These studies, also known as GWAS (Genome Wide Association Study), are based on the assumption that genetic susceptibility is caused by the interaction of certain common genetic variants. In fact, being multifactorial diseases frequent diseases (just think of diabetes mellitus type II or ischemic heart disease), the underlying assumption is: frequent disease, FREQUENT variant. NGS has revolutionized this research field too, with genome/exome sequencing having already replaced genome/exome array, which allow the screening of just a few variants (exome sequencing, because of its affordability compared to genome sequencing, is actually the most used one). Of note, the philosophy underlying the use of genome/exome sequencing in multiple disease research studies, is exactly the opposite of genome/exome array. It is: frequent disease, RARE variant (i.e. that susceptibility to multifactorial disease is more about the interaction of rare rather than frequent variants).
3. ROUTINE DIAGNOSIS
Exome sequencing is certainly becoming very useful in routine diagnosis too. Sequencing the entire coding region can be economically advantageous (especially in respect to certain Sanger sequencing analyses) and offers the chance to identify the genetic cause even when clinical confirmation is poor or not specific enough to limit the diagnostic suspicion to a number of possible disorders.
