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Open Access Highly Accessed Methodology article

AluScan: a method for genome-wide scanning of sequence and structure variations in the human genome

Lingling Mei1, Xiaofan Ding1, Shui-Ying Tsang1, Frank W Pun1, Siu-Kin Ng1, Jianfeng Yang1, Cunyou Zhao1, Dezhi Li2, Weiqing Wan23, Chi-Hung Yu45, Tze-Ching Tan45, Wai-Sang Poon56, Gilberto Ka-Kit Leung57, Ho-Keung Ng58, Liwei Zhang23* and Hong Xue135*

Author Affiliations

1 Division of Life Science and Applied Genomics Centre, Hong Kong University of Science and Technology, 1 University Road, Clear Water Bay, Kowloon, Hong Kong, China

2 Department of Neurosurgery, Beijing Tiantan Hospital, 6 Tiantan Xili, Dongcheng District, Capital Medical University, Beijing, 100050, China

3 Chinese Cancer Genome Consortium, Beijing Genome Institute Shenzhen, 11 Beishan Industrial Zone, Yantian District, Shenzhen, 518083, China

4 Department of Neurosurgery, Queen Elizabeth Hospital, 30 Gascoigne Road, Kowloon, Hong Kong, China

5 Brain Cancer Genome Consortium - Hong Kong, Applied Genomics Center, Hong Kong University of Science and Technology, 1 University Road, Clear Water Bay, Kowloon, Hong Kong, China

6 Division of Neurosurgery, Department of Surgery, Prince of Wales Hospital, Chinese University of Hong Kong, 30-32 Ngan Shing Street, Sha Tin, Hong Kong, China

7 Division of Neurosurgery, Department of Surgery, Li Ka Shing Faculty of Medicine, University of Hong Kong, Queen Mary Hospital, 102 Pokfulam Road, Hong Kong, China

8 Department of Anatomical and Cellular Pathology, Prince of Wales Hospital, Chinese University of Hong Kong, 30-32 Ngan Shing Street, Sha Tin, Hong Kong, China

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BMC Genomics 2011, 12:564  doi:10.1186/1471-2164-12-564

Published: 17 November 2011

Abstract

Background

To complement next-generation sequencing technologies, there is a pressing need for efficient pre-sequencing capture methods with reduced costs and DNA requirement. The Alu family of short interspersed nucleotide elements is the most abundant type of transposable elements in the human genome and a recognized source of genome instability. With over one million Alu elements distributed throughout the genome, they are well positioned to facilitate genome-wide sequence amplification and capture of regions likely to harbor genetic variation hotspots of biological relevance.

Results

Here we report on the use of inter-Alu PCR with an enhanced range of amplicons in conjunction with next-generation sequencing to generate an Alu-anchored scan, or 'AluScan', of DNA sequences between Alu transposons, where Alu consensus sequence-based 'H-type' PCR primers that elongate outward from the head of an Alu element are combined with 'T-type' primers elongating from the poly-A containing tail to achieve huge amplicon range. To illustrate the method, glioma DNA was compared with white blood cell control DNA of the same patient by means of AluScan. The over 10 Mb sequences obtained, derived from more than 8,000 genes spread over all the chromosomes, revealed a highly reproducible capture of genomic sequences enriched in genic sequences and cancer candidate gene regions. Requiring only sub-micrograms of sample DNA, the power of AluScan as a discovery tool for genetic variations was demonstrated by the identification of 357 instances of loss of heterozygosity, 341 somatic indels, 274 somatic SNVs, and seven potential somatic SNV hotspots between control and glioma DNA.

Conclusions

AluScan, implemented with just a small number of H-type and T-type inter-Alu PCR primers, provides an effective capture of a diversity of genome-wide sequences for analysis. The method, by enabling an examination of gene-enriched regions containing exons, introns, and intergenic sequences with modest capture and sequencing costs, computation workload and DNA sample requirement is particularly well suited for accelerating the discovery of somatic mutations, as well as analysis of disease-predisposing germline polymorphisms, by making possible the comparative genome-wide scanning of DNA sequences from large human cohorts.