A multiplex single-base extension protocol for genotyping Cdx2, FokI, BsmI, ApaI, and TaqI polymorphisms of the vitamin D receptor gene.
The well-described role of the vitamin D endocrine system in bone metabolism makes its receptor a widely investigated candidate gene in association studies looking for the genetic basis of complex bone-related phenotypes. Most association studies genotype five polymorphic sites along the gene using PCR-RFLP and allele-specific amplification methods, which may not be the better choice in large case/control or cross-sectional studies. In this case, genotyping SNPs in parallel and using automated allele-calling methods are important to decrease genotyping errors due to manual data handling and save sample in cases where the amount of DNA is limited. The aim of this study was to present a straightforward method based on multiplex PCR amplification followed by multiplex single-base extension as a simple way to genotype five vitamin D receptor gene polymorphisms in parallel, which may be implemented in medium- to large-scale case/control or cross-sectional studies. The results regarding method feasibility and optimization are presented by genotyping eight paternity trios and seven samples of Brazilian postmenopausal women who took part in an ongoing association study carried out by members of our group.
Lins
T C L
TC
Pós-Graduação em Ciências Genômicas e Biotecnologia, Universidade Católica de Brasília, Brasília, DF, Brasil.
Nogueira
L R
LR
Lima
R M
RM
Gentil
P
P
Oliveira
R J
RJ
Pereira
R W
RW
D001938:Brazil; D002648:Child; D019075:Electrophoresis, Capillary; D005260:Female; D005838:Genotype; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D017698:Postmenopause; D018167:Receptors, Calcitriol / Q000235:genetics*
D018167:Receptors, Calcitriol