Gene Hunting Approaches through the Combination of Linkage Analysis with Whole-Exome Sequencing in Mendelian Diseases: From Darwin to the Present Day

<b><i>Background:</i></b> In the context of medical genetics, gene hunting is the process of identifying and functionally characterizing genes or genetic variations that contribute to disease phenotypes. In this review, we would like to summarize gene hunting process in terms of historical aspects from Darwin to now. For this purpose, different approaches and recent developments will be detailed. <b><i>Summary:</i></b> Linkage analysis and association studies are the most common methods in use for explaining the genetic background of hereditary diseases and disorders. Although linkage analysis is a relatively old approach, it is still a powerful method to detect disease-causing rare variants using family-based data, particularly for consanguineous marriages. As is known that, consanguineous marriages or endogamy poses a social problem in developing countries, however, this same condition also provides a unique opportunity for scientists to identify and characterize pathogenic variants. The rapid advancements in sequencing technologies and their parallel implementation together with linkage analyses now allow us to identify the candidate variants related to diseases in a relatively short time. Furthermore, we can now go one step further and functionally characterize the causative variant through in vitro and in vivo studies and unveil the variant-phenotype relationships on a molecular level more robustly. <b><i>Key Messages:</i></b> Herein, we suggest that the combined analysis of linkage and exome analysis is a powerful and precise tool to diagnose clinically rare and recessively inherited conditions.

The History of Gene Hunting in Hereditary Spinocerebellar Degeneration: Lessons From the Past and Future Perspectives

Hereditary spinocerebellar degeneration (SCD) encompasses an expanding list of rare diseases with a broad clinical and genetic heterogeneity, complicating their diagnosis and management in daily clinical practice. Correct diagnosis is a pillar for precision medicine, a branch of medicine that promises to flourish with the progressive improvements in studying the human genome. Discovering the genes causing novel Mendelian phenotypes contributes to precision medicine by diagnosing subsets of patients with previously undiagnosed conditions, guiding the management of these patients and their families, and enabling the discovery of more causes of Mendelian diseases. This new knowledge provides insight into the biological processes involved in health and disease, including the more common complex disorders. This review discusses the evolution of the clinical and genetic approaches used to diagnose hereditary SCD and the potential of new tools for future discoveries.

Fox Hunting in Wild Apples: Searching for Novel Genes in Malus Sieversii

Malus sieversii is considered the progenitor of modern apple (Malus pumila) cultivars and to represent a valuable source of genetic diversity. Despite the importance of M. sieversii as a source of disease resistance, stress tolerance, and novel fruit traits, little is known about gene function and diversity in M. sieversii. Notably, a publicly annotated genome sequence for this species is not available. In the current study, the FOX (Full-length cDNA OvereXpressing) gene hunting system was used to construct a library of transgenic lines of Arabidopsis in which each transgenic line overexpresses a full-length gene obtained from a cDNA library of the PI619283 accession of M. sieversii. The cDNA library was constructed from mRNA obtained from bark tissues collected in late fall–early winter, a time at which many abiotic stress-adaptative genes are expressed. Over 4000 apple FOX Arabidopsis lines have been established from the pool of transgenic seeds and cDNA inserts corresponding to various Gene Ontology (GO) categories have been identified. A total of 160 inserts appear to be novel, with no or limited homology to M. pumila, Arabidopsis, or poplar. Over 1300 lines have also been screened for freezing resistance. The constructed library of transgenic lines provides a valuable genetic resource for exploring gene function and diversity in Malus sieversii. Notably, no such library of t-DNA lines currently exists for any Malus species.