title: A novel approach for copy number variation analysis by combining multiplex PCR with matrix-assisted laser desorption ionization time-of-flight mass spectrometry.
abstract: With the increasing interest in copy number variation as it pertains to human genomic variation, common phenotypes, and disease susceptibility, there is a pressing need for methods to accurately identify copy number. In this study, we developed a simple approach that combines multiplex PCR with matrix-assisted laser desorption ionization time-of-flight mass spectrometry for submicroscopic copy number variation detection. Two pairs of primers were used to simultaneously amplify query and endogenous control regions in the same reaction. Using a base extension reaction, the two amplicons were then distinguished and quantified in a mass spectrometry map. The peak ratio between the test region and the endogenous control region was manually calculated. The relative copy number could be determined by comparing the peak ratio between the test and control samples. This method generated a copy number measurement comparable to those produced by two other commonly used methods - multiplex ligation-dependent probe amplification and quantitative real-time PCR. Furthermore, it can discriminate a wide range of copy numbers. With a typical 384-format SpectroCHIP, at least six loci on 384 samples can be analyzed simultaneously in a hexaplex assay, making this assay adaptable for high throughput, and potentially applicable for large-scale association studies.
authors: Capital Institute of Pediatrics, Beijing 100020, China.
authors: Yonghui
authors: Y
authors: Gao
authors: Xiaoli
authors: X
authors: Chen
authors: Jianhua
authors: J
authors: Wang
authors: Shaofang
authors: S
authors: Shangguan
authors: Yaohua
authors: Y
authors: Dai
authors: Ting
authors: T
authors: Zhang
authors: Junling
authors: J
authors: Liu
mesh_terms: D001483:Base Sequence; D002648:Child; D056915:DNA Copy Number Variations; D004252:DNA Mutational Analysis / Q000379:methods*; D005260:Female; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D008297:Male; D046208:Microchip Analytical Procedures; D008954:Models, Biological; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*