A multiplex single nucleotide polymorphism genotyping method using ligase‐based mismatch discrimination and CE‐SSCP |
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| Authors: | Woong Choi Gi Won Shin Hee Sung Hwang Seung Pil Pack Gyu Yong Jung Gyoo Yeol Jung |
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| Affiliation: | 1. School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, Pohang, Gyeongbuk, Korea;2. Institute of Environmental and Energy Technology, Pohang University of Science and Technology, Pohang, Gyeongbuk, Korea;3. Department of Biotechnology and Bioinformatics, Korea University, Jochiwon, Sejong, Korea;4. Department of Plastic Surgery, College of Medicine, Dongguk University, Gyeongju, Gyeongbuk, Korea;5. Department of Chemical Engineering, Pohang University of Science and Technology, Pohang, Gyeongbuk, Korea |
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| Abstract: | Accuracy, simplicity, and cost‐effectiveness are the most important criteria for a genotyping method for SNPs compatible with clinical use. One method developed for SNP genotyping, ligase‐based discrimination, is considered the simplest for clinical diagnosis. However, multiplex assays using this method are limited by the detection method. Although CE has been introduced as an alternative to error prone microarray‐based detection, the design process and multiplex assay procedure are complicated because of the DNA size‐dependent separation principle. In this study, we developed a simple and accurate multiplex genotyping method using reaction condition‐optimized ligation and high‐resolution CE‐based SSCP. With this high‐resolution CE‐SSCP system, we are able to use similar‐sized probes, thereby eliminating the complex probe design step and simplifying the optimization process. We found that this method could accurately discriminate single‐base mismatches in SNPs of the tp53 gene, used as targets for multiplex detection. |
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| Keywords: | CE‐SSCP Diagnostic assay Ligase‐based genotyping assay Multiplex analysis SNP |
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