Highly multiplex and sensitive SNP genotyping method using a three‐color fluorescence‐labeled ligase detection reaction coupled with conformation‐sensitive CE

For the development of clinically useful genotyping methods for SNPs, accuracy, simplicity, sensitivity, and cost‐effectiveness are the most important criteria. Among the methods currently being developed for SNP genotyping technology, the ligation‐dependent method is considered the simplest for cli...

Full description

Saved in:
Bibliographic Details
Published in:Electrophoresis 2017-02, Vol.38 (3-4), p.513-520
Main Authors: Choi, Woong, Jung, Gyoo Yeol
Format: Article
Language:English
Subjects:
Amplification
Assaying
CE‐SSCP
Color
Criteria
Denaturation
Deoxyribonucleic acid
Design analysis
Diagnosis
DNA
Electrophoresis, Capillary - methods
Error detection
Fluorescence
Fluorescent Dyes - chemistry
Genes
Genotyping Techniques - methods
High resolution
Humans
Ligase detection reaction
Ligases - analysis
Ligases - chemistry
Ligases - metabolism
Multiplex detection
Multiplexing
Polymorphism, Single Nucleotide - genetics
Polymorphism, Single-Stranded Conformational - genetics
Sensitivity analysis
Separation
SNP
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:For the development of clinically useful genotyping methods for SNPs, accuracy, simplicity, sensitivity, and cost‐effectiveness are the most important criteria. Among the methods currently being developed for SNP genotyping technology, the ligation‐dependent method is considered the simplest for clinical diagnosis. However, sensitivity is not guaranteed by the ligation reaction alone, and analysis of multiple targets is limited by the detection method. Although CE is an attractive alternative to error‐prone hybridization‐based detection, the multiplex assay process is complicated because of the size‐based DNA separation principle. In this study, we employed the ligase detection reaction coupled with high‐resolution CE‐SSCP to develop an accurate, sensitive, and simple multiplex genotyping method. Ligase detection reaction could amplify ligated products through recurrence of denaturation and ligation reaction, and SSCP could separate these products according to each different structure conformation without size variation. Thus, simple and sensitive SNP analysis can be performed using this method involving the use of similar‐sized probes, without complex probe design steps. We found that this method could not only accurately discriminate base mismatches but also quantitatively detect 37 SNPs of the tp53 gene, which are used as targets in multiplex analysis, using three‐color fluorescence‐labeled probes.
ISSN:0173-0835
1522-2683
DOI:10.1002/elps.201600369