detection of chromosomal structural alterations in single cells by snp arrays a systematic survey of amplification bias and optimized workflow检测单个细胞的染色体结构改变snp阵列系统放大偏差的调查和优化工作流程.pdfVIP

Detection of Chromosomal Structural Alterations in Single Cells by SNP Arrays: A Systematic Survey of Amplification Bias and Optimized Workflow 1 1 1 1 2 2 2 1 Kazuya Iwamoto *, Miki Bundo , Junko Ueda , Yoko Nakano , Wataru Ukai , Eri Hashimoto , Toshikazu Saito , Tadafumi Kato 1 Laboratory for Molecular Dynamics of Mental Disorders, RIKEN Brain Science Institute, Wako, Saitama, Japan, 2 Department of Neuropsychiatry, Sapporo Medical University, Chuo-Ku, Sapporo, Japan Background. In single-cell human genome analysis using whole-genome amplified product, a strong amplification bias involving allele dropout and preferential amplification hampers the quality of results. Using an oligonucleotide single nucleotide polymorphism (SNP) array, we systematically examined the nature of this amplification bias, including frequency, degree, and preference for genomic location, and we assessed the effects of this amplification bias on subsequent genotype and chromosomal copy number analyses. Methodology/Principal Findings. We found a large variability in amplification bias among the amplified products obtained by multiple displacement amplification (MDA), and this bias had a severe effect on the genotype and chromosomal copy number analyses. We established optimal experimental conditions for pre-screening for high-quality amplified products, processing array data, and analyzing chromosomal structural alterations. Using this optimized protocol, we successfully detected previously unidentified chromosomal structural alterations in single cells from a lymphoblastoid cell line. These alterations were subsequently confirmed by karyotype analysis. In addition, we successfully obtained reproducible chromosomal