Ultra-short reads and non-model species: Exploring the complexities of next generation sequence assembly and SNP discovery in the absence of a reference genome.

How practical is gene and SNP discovery in a non-model species using ultra-short read sequences? Next generation sequencing technologies are being applied to an increasing number of species with no reference genome. For labs working with non-model species, the cost, availability of existing genetic resources, sequence complexity, and the planned method of assembly must all be considered when selecting a sequencing platform. Our goal is to examine the feasibility and optimal methodology for SNP and gene discovery in the sockeye salmon (Oncorhynchus nerka), using ultra-short read sequences. SOLiD short reads (up to 50bp) were generated from single and pooled tissue transcriptome libraries from each of ten O. nerka individuals. The individuals were from five distinct populations, from the Wood River Lakes and Southeast Alaska. As no reference genome is available for O. nerka, the SOLiD sequence reads were aligned to publicly available EST references sequences from O. nerka and two closely related species, rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar). Additionally, de novo assembly of the SOLiD data was carried out using the CLC NGS Cell. The results from each reference assembly were compared across species, as well as to the results of de novo assembly, to determine the feasibility and optimal methodology for SNP discovery using ultra-short reads in a non-model species.