Single-stranded conformation polymorphism analysis of Vitamin D receptor gene by capillary electrophoresis with laser-induced fluorescence detection

A highly sensitive single-stranded conformation polymorphism method for analysis of Vitamin D receptor (VDR) gene polymorphism was developed employing laser-induced fluorescence capillary electrophoresis (LIF-CE). LIF-CE was conducted utilizing a linear polyacrylamide solution as entangled polymer and acridine orange as a fluorescent dye of single-stranded DNA. Effect of acridine orange, size of PCR product and running temperature were investigated by LIF-CE in order to analyze the two polymorphisms of the allelic variation of the Bsm I and Taq I sites in intron 8 and exon 9, respectively, of the VDR gene. The developed method was simple, rapid (< 16 min for Taq I type, < 20 min for Bsm I type) and highly sensitive for VDR gene polymorphism. VDR gene polymorphism in 32 subjects was determined by the proposed method. The results were consistent with those obtained by restriction fragment-length polymorphism (RFLP) analysis using gel electrophoresis. The proposed method can be employed among the various VDR gene polymorphism analyses related to osteoporosis. (C) 2001 Elsevier Science B.V. All rights reserved.