AluScan sequencing and its application to cancer genomics

Abstract

Background While next generation sequencing (NGS) has enabled massive parallel analysis of different human genomes, strategies are required to maximize its signal‐to‐noise ratio in the data generated and its accessibility for clinical settings. While whole exome sequence (WES) can capture subsets of whole genome sequences (WGS), thereby reducing cost and time of analysis, it is inapplicable to non‐protein coding sequences. Pre‐sequencing capture methods are therefore needed to optimize the collection of genome‐wide sequence information based on minimal deoxyribonucleic acid (DNA) samples for a broad variety of applications. Aims Because retrotransposons of the Alu family are widespread in the human genome, especially in gene dense regions, their consensus sequences have been employed to design polymerase chain reaction (PCR) primers capable of the simultaneous amplification of randomly distributed inter‐Alu sequences across the genome, in the form of an AluScan sequencing platform. This combination of multiplex inter‐Alu PCR with NGS have yielded huge numbers of inter‐Alu sequences in all regions of the human genome that expedite enquiry into the roles of different sequence and structural variations using only submicrograms of template genomic DNA. Accordingly, the aim of this study was to examine how the AluScan platform could be employed to analyze the somatic and germline single‐nucleotide‐variations (SNVs) and copy‐number‐variations (CNVs) related to cancers, in order to delineate some of the occurences and concequences of such mutations. Methodologies NGS has undergone rapid development and expansion in recent years, and WGS reprecents a pivotal contributor to the deepening understanding of disease‐genome relationships. However, the considerable workload associated with WGS has limited large scale investigations on a population basis. While WES analysis captures a subset of total sequences and thus economizes on workload and costs, its application is limited to the genic regions of the genome. Therefore there is a need to have a method that could economize without extensive loss of genomic sequences, such as the simultaneous amplification of a huge number of inter‐Alu seqences on the AluScan platform which is rendered efficient by the huge number of Alu retrotransposons distributed over genic as well as non‐genic regions of the genome, and examined in the present study. Result and Conclusion The utility of the AluScan method was illustrated by its usage in delineating the central role of interstitial loss‐of‐heterozygosity (LOH) mutations in cancers, the widespread occurence of forward‐reverse mutation cycles between different stages of cancer development, prognosis of liver cancers based on their relative abundances in LOHs and somatic copy‐number‐variations (CNVs), and prediction of the susceptibility of a genome to cancers based on reccurent germline CNVs. Moreover, owing to the concentration of hotspots and hotspot clusters of genomic features in the vicinities of Alu and other retrotransposons,...