title: Identification of novel SNPs by next-generation sequencing of the genomic region containing the APC gene in colorectal cancer patients in China.
abstract: We described an approach of identifying single nucleotide polymorphisms (SNPs) in complete genomic regions of key genes including promoters, exons, introns, and downstream sequences by combining long-range polymerase chain reaction (PCR) or NimbleGen sequence capture with next-generation sequencing. Using the adenomatous polyposis coli (APC) gene as an example, we identified 210 highly reliable SNPs by next-generation sequencing analysis program MAQ and Samtools, of which 69 were novel ones, in the 123-kb APC genomic region in 27 pair of colorectal cancers and normal adjacent tissues. We confirmed all of the eight randomly selected high-quality SNPs by allele-specific PCR, suggesting that our false discovery rate is negligible. We identified 11 SNPs in the exonic region, including one novel SNP that was not previously reported. Although 10 of them are synonymous, they were predicted to affect splicing by creating or removing exonic splicing enhancers or exonic splicing silencers. We also identified seven SNPs in the upstream region of the APC gene, three of which were only identified in the cancer tissues. Six of these upstream SNPs were predicted to affect transcription factor binding. We also observed that long-range PCR was better in capturing GC-rich regions than the NimbleGen sequence capture technique.
authors.affiliation: Systems Biology Division, Zhejiang-California International Nanosystems Institute (ZCNI), Zhejiang University, Hangzhou, People's Republic of China.
authors.forename: Yin
authors.initials: Y
authors.lastname: Cheng
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.forename: Jiaofang
authors.initials: J
authors.lastname: Shao
authors.forename: Qiyun
authors.initials: Q
authors.lastname: Chen
authors.forename: Fan
authors.initials: F
authors.lastname: Mo
authors.forename: Liang
authors.initials: L
authors.lastname: Ma
authors.forename: Xu
authors.initials: X
authors.lastname: Han
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.forename: Chen
authors.initials: C
authors.lastname: Chen
authors.forename: Cixiong
authors.initials: C
authors.lastname: Zhang
authors.forename: Shuyong
authors.initials: S
authors.lastname: Lin
authors.forename: Jiekai
authors.initials: J
authors.lastname: Yu
authors.forename: Shu
authors.initials: S
authors.lastname: Zheng
authors.forename: Sheng-Cai
authors.initials: SC
authors.lastname: Lin
authors.forename: Biaoyang
authors.initials: B
authors.lastname: Lin
mesh_terms: D001483:Base Sequence; D002681:China; D015179:Colorectal Neoplasms / Q000235:genetics*; D017491:Genes, APC; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods*; D012984:Software