Candidate gene testing in clinical cohort studies with multiplexed genotyping and mass spectrometry

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Item Type:	Article
Title:	Candidate gene testing in clinical cohort studies with multiplexed genotyping and mass spectrometry
Creators Name:	Ashley, S.E., Meyer, B.A., Ellis, J.A. and Martino, D.J.
Abstract:	Complex diseases are often underpinned by multiple common genetic variants that contribute to disease susceptibility. Here, we describe a cost-effective tag single nucleotide polymorphism (SNP) approach using a multiplexed genotyping assay with mass spectrometry, to investigate gene pathway associations in clinical cohorts. We investigate the food allergy candidate locus Interleukin13 (IL13) as an example. This method efficiently maximizes the coverage by taking advantage of shared linkage disequilibrium (LD) within a region. Selected LD SNPs are then designed into a multiplexed assay, enabling up to 40 different SNPs to be analyzed simultaneously, boosting cost-effectiveness. Polymerase chain reaction (PCR) is used to amplify the target loci, followed by single nucleotide extension, and the amplicons are then measured using matrix-assisted laser desorption/ionization-time of flight(MALDI-TOF) mass spectrometry. The raw output is analyzed with the genotype calling software, using stringent quality control definitions and cut-offs, and high probability genotypes are determined and output for data analysis.
Keywords:	Cohort Studies, Genetic Testing, Genotype, Mass Spectrometry
Source:	Journal of Visualized Experiments
ISSN:	1940-087X
Publisher:	JoVE
Number:	136
Page Range:	e57601
Date:	21 June 2018
Additional Information:	This article will also be available in Open Access at PubMed Central with a delayed release (embargo) on June 21, 2020.
Official Publication:	https://doi.org/10.3791/57601
External Fulltext:	View full text on PubMed Central
PubMed:	View item in PubMed

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