Polymorphism in IRF6 and orofacial clefting: searching for a fast detection technique

CHIRIOTTO TS; CÓRDOBA EE; DE GANZÓ AF; DÉRRICI R; ABRILE GW; CHEROKI C

Viña del Mar

Congreso; Congreso Latinoamiricano de Genética (ALAG); 2010

Universidad Andrés Bello

Nonsyndromic cleft lip with or without cleft palate (CL/P NS) is a complex disorder in the development of the orofacial structures. The incidence of these defect is 1-2:1000 live births worldwide. The IRF6 gene is one of the many orofacial cleft locus involved in this disorder. Point mutations in this gene have been related to disruptions in the facial development of normal mice embryos, thus, been related to the increasing risk for the pathology in humans. In this study, we search for the optimization of a detection technique for one specific SNP, a nonsynonymous substitution, G820A, located in the exon 7 of the IRF6 gene. For that reason, members of an affected pedigree were analyzed to develop a G820A detection technique. Their DNA was obtained from peripheral blood or oral swabs using CTAB or TEC-SDS techniques, respectively. Specific primers to amplify the whole exon (586bp) were designed and PCR conditions were optimized. PCR products were digested with Mbol enzyme and the digested products were visualized in 2% agar gel electrophoresis. Amplified products were also sequenced to confirm the genotypes obtained (G/G, G/A, A/A). We obtained a detection technique for the exon 7 SNP witch is fast and simple and can be apply in any molecular biology laboratory. Our technique is able to establish the genotypes of any individual (normal or affected) inside the pedigree, providing useful information in terms of genetic counselling