title: Simple Method to Genotype the ACTN3 r577x Polymorphism.
abstract: The alpha-actinin-3 r577x polymorphism (rs1815739) is one of the most important polymorphisms associated with athletic performance. This single-nucleotide mutation leads to a premature stop codon, resulting in a nonfunctional protein product. The presence of the dominant R allele is associated with full power skeletal muscle contraction. Homozygosity for the X allele is correlated with more efficient energy disposure. Restriction fragment length polymorphism and real-time polymerase chain reaction (PCR) are the standard methods used to genotype this polymorphism, but they are expensive and require special equipments. Here, we present a simple and cost-efficient method to genotype the ACTN3 r577x polymorphism by a single PCR. External primers yield a 690-bp product that indicates the template quality. Internal primers produce a 413-bp product if the R allele is present and a 318-bp product if the X allele is present. Our four-primer genotyping PCR was validated by the standard real-time PCR, generally used to genotype this single-nucleotide polymorphism, demonstrating the accuracy of this method. This protocol is perfect for small- or large-scale cohort genotyping of the ACTN3 r577x polymorphism.
authors.affiliation: 1 Laboratory of Nutrigenomics and Metabology, Federal University of Pelotas , Pelotas, Rio Grande do Sul, Brazil .
authors.forename: Ines
authors.initials: I
authors.lastname: Schadock
authors.forename: Augusto
authors.initials: A
authors.lastname: Schneider
authors.forename: Elton Dias
authors.initials: ED
authors.lastname: Silva
authors.forename: Marcia Rubia Duarte
authors.initials: MR
authors.lastname: Buchweitz
authors.forename: Marcio Nunes
authors.initials: MN
authors.lastname: Correa
authors.forename: Joao Bosco
authors.initials: JB
authors.lastname: Pesquero
authors.forename: Edgar Julian
authors.initials: EJ
authors.lastname: Paredes-Gamero
authors.forename: Ronaldo Carvalho
authors.initials: RC
authors.lastname: Araujo
authors.forename: Carlos Castilho
authors.initials: CC
authors.lastname: Barros
mesh_terms: D000185:Actinin / Q000235:genetics*; D000199:Actins / Q000235:genetics; D054874:Athletic Performance / Q000502:physiology; D001483:Base Sequence; D017931:DNA Primers; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide
chemical_list: C118456:ACTN3 protein, human; D000199:Actins; D017931:DNA Primers; D000185:Actinin