Open AccessHigh‐Resolution Melting Analysis as a Developed Method for Genotyping the PD Susceptibility Loci in LRRK2 Gene
Author(s) -
Jiang Enzhu,
Li Fengrui,
Jing Chenchen,
Li Pei,
Cui Honggang,
Wang Baojie,
Ding Mei,
Pang Hao
Publication year - 2015
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.21769
Subject(s) - high resolution melt , genotyping , single nucleotide polymorphism , amplicon , genetics , biology , snp genotyping , melting curve analysis , genotype , multiplex , gene , microbiology and biotechnology , polymerase chain reaction
Background Single‐nucleotide polymorphisms (SNPs) have been reported as a highly relevant point for the mechanisms of Parkinson's disease (PD). The invention of saturating dye makes it possible to identify heteroduplex DNA without redistribution during melting, which allows using high‐resolution melting (HRM) to detect SNPs. However, the HRM analysis for detection of those SNPs associated with PD was rarely applied. Methods Two SNPs, G2385R and R1628P , located in leucine‐rich repeat kinase 2 ( LRRK2 ) gene were individually and multiplexedly genotyped using HRM analysis. The sequence variant observed in unexpected HRM curves was confirmed by DNA sequencing. Results HRM analysis identified successfully all genotypes both on R1628P and G2385R loci. The unexpected HRM curves appeared in R1628P amplicon generated from combinations of R1628P and rs11176013 loci. A multiplexed HRM assay that genotyped R1628P , rs11176013, and G2385R loci was efficiently established. Conclusions The present HRM assay is a reliable and rapid method for genotyping R1628P and G2385R loci in LRRK2 gene, and multiplex HRM analysis results in high throughput and has the potential to facilitate a wide range of genotyping studies on PD susceptibility genes.