title: Identification of RFLP G6PD mutations by using microcapillary electrophoretic chips (Experion).
abstract: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is one of the most well-known human genetic defects, identified in more than 400 million individuals in the world. To date, no commercial kits are available for the mutation screening of this disease. Seventy G6PD-deficient Italian individuals admitted to the Laboratory of Clinical Molecular Biology of Hospital "Agostino Gemelli" of Rome were screened for the most frequent Italian mutations, by means of allele-specific PCR, followed by restriction fragment length electrophoresis. The present study compares two techniques for the identification of restriction patterns: agarose gel electrophoresis versus Experion system. When the first screening was negative, the entire G6PD gene was sequenced using the ABI 3100 Avant Instrumentation. The G6PD variants identified and their frequencies were the following: G6PD Mediterranean (75.7%), G6PD Seattle (7.1%), G6PD A(-) 202 + 376 (7.1%), and G6PD Cassano (2.8%). In addition, we identified by direct sequencing two new mutations, namely Buenos Aires and Rignano. With the Experion method, the size band determination was more accurate than that obtained by gel electrophoresis. The Experion system resulted as a valid, easy, and reproducible diagnostic method for the screening of G6PD mutation as compared with the agarose electrophoretic analysis.
authors.affiliation: Laboratory of Clinical Molecular Biology, Institute of Biochemistry and Clinical Biochemistry, Catholic University, Rome, Italy.
authors.forename: Angelo
authors.initials: A
authors.lastname: Minucci
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Delibato
authors.forename: Massimo
authors.initials: M
authors.lastname: Castagnola
authors.forename: Paola
authors.initials: P
authors.lastname: Concolino
authors.forename: Franco
authors.initials: F
authors.lastname: Ameglio
authors.forename: Cecilia
authors.initials: C
authors.lastname: Zuppi
authors.forename: Bruno
authors.initials: B
authors.lastname: Giardina
authors.forename: Ettore
authors.initials: E
authors.lastname: Capoluongo
mesh_terms: D004252:DNA Mutational Analysis / Q000295:instrumentation / Q000379:methods; D004587:Electrophoresis, Agar Gel / Q000295:instrumentation / Q000379:methods; D046209:Electrophoresis, Microchip / Q000295:instrumentation / Q000379:methods*; D005260:Female; D005787:Gene Frequency; D005820:Genetic Testing / Q000379:methods; D005954:Glucosephosphate Dehydrogenase / Q000032:analysis* / Q000235:genetics*; D006801:Humans; D008297:Male; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D005954:Glucosephosphate Dehydrogenase