Teaching molecular genetics: chapter 4—positional cloning of genetic disorders
Positional cloning is the approach of choice for the identification of genetic mutations underlying the pathological development of diseases with simple Mendelian inheritance. It consists of different consecutive steps, starting with recruitment of patients and DNA collection, that are critical to the overall process. A genetic analysis of the enrolled patients and their families is performed, based on genetic recombination frequencies generated by meiotic cross-overs and on genome-wide molecular studies, to define a critical DNA region of interest. This analysis culminates in a statistical estimate of the probability that disease features may segregate in the families independently or in association with specific molecular markers located in known regions. In this latter case, a marker can be defined as being linked to the disease manifestations. The genetic markers define an interval that is a function of their recombination frequencies with the disease, in which the disease gene is localised. The identification and characterisation of chromosome abnormalities as translocations, deletions and duplications by classical cytogenetic methods or by the newly developed microarray-based comparative genomic hybridisation (array CGH) technique may define extensions and borders of the genomic regions involved. The step following the definition of a critical genomic region is the identification of candidate genes that is based on the analysis of available databases from genome browsers. Positional cloning culminates in the identification of the causative gene mutation, and the definition of its functional role in the pathogenesis of the disorder, by the use of cell-based or animal-based experiments. More often, positional cloning ends with the generation of mice with homologous mutations reproducing the human clinical phenotype. Altogether, positional cloning has represented a fundamental step in the research on genetic renal disorders, leading to the definition of several disease mechanisms and allowing a proper diagnostic approach to many conditions.
KeywordsPositional cloning Linkage analysis Mutation screening Gene functional analysis Mendelian renal disease
Data were critically discussed with Prof. Rosanna Gusmano, and we acknowledge her role. The manuscript was revised by Miss Capurro.
- 1.Strachan T, Read A (2003) Human molecular genetics, 3rd edn. Garland, LondonGoogle Scholar
- 12.Hinkes B, Wiggins RC, Gbadegesin R, Vlangos CN, Seelow D, Nurnberg G, Garg P, Verma R, Chaib H, Hoskins BE, Ashraf S, Becker C, Hennies HC, Goyal M, Wharram BL, Schachter AD, Mudumana S, Drummond I, Kerjaschki D, Waldherr R, Dietrich A, Ozaltin F, Bakkaloglu A, Cleper R, Basel-Vanagaite L, Pohl M, Griebel M, Tsygin AN, Soylu A, Muller D, Sorli CS, Bunney TD, Katan M, Liu J, Attanasio M, O’Toole JF, Hasselbacher K, Mucha B, Otto EA, Airik R, Kispert A, Kelley GG, Smrcka AV, Gudermann T, Holzman LB, Nurnberg P, Hildebrandt F (2006) Positional cloning uncovers mutations in PLCE1 responsible for a nephrotic syndrome variant that may be reversible. Nat Genet 38:1397–1405PubMedCrossRefGoogle Scholar