Abstract
Many techniques for the study of complex populations provide either specific information on a small number of variants or general information on the entire population. Here we describe a powerful new technique for elucidating mutation frequencies at each genomic position in a complex population. This single base extension (SBE) based microarray platform was designed and optimized using poliovirus as the target genotype, but can be easily adapted to assay populations derived from any organism. The sensitivity of the method was demonstrated by accurate and consistent readouts from a controlled population of mutant genotypes. We subsequently deployed the technique to investigate the effects of the nucleotide analog ribavirin on a typical poliovirus population through two rounds of passage. Our results show that this economical platform can be used to investigate dynamic changes occurring at frequencies below 1% within a complex nucleic acid population. Given that many key aspects of the study and treatment of disease are intimately linked to population-level genomic diversity, our SBE-based technique provides a scalable and cost-effective complement to both traditional and next generation sequencing methodologies.
Highlights
The identification of specific genetic changes responsible for molecular, cellular, and organismal phenotypes is central to many biological questions
We demonstrate the performance of our novel mutation distribution analysis of populations (MDAP) microarray by characterizing the mutant spectrum of a poliovirus population subjected to a mutagenic drug
We developed a single base extension microarray platform with the goal of determining individual nucleotide frequencies at each genomic position within a complex viral population
Summary
The identification of specific genetic changes responsible for molecular, cellular, and organismal phenotypes is central to many biological questions. A phenotype is linked to either the responsible gene or a segregating polymorphism. The power of this approach has recently increased with the development of high-density single nucleotide polymorphism maps in several organisms [1,2,3]. Many questions require quantification of allele frequency or genetic diversity at the population level. Genetic association studies of complex traits often rely on measurements of several independently segregating single nucleotide polymorphisms in diverse populations [4]. Genetic rearrangements and somatic hypermutation lead to extensive diversity within B and T cell populations [6,7,8]
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