A Research Workflow for Development of Single-Copy Nuclear Loci and High-throughput Targeted Enrichment for Two or More Phylogenetic Studies.
Jordon-Thaden, I., Uribe-Convers, S., Godden, G.T., Srikar, C., Tank, D.C., Martine, C.T.Many laboratories in evolutionary biology are interested in reducing research expenses by coordinating laboratory efforts and combining samples from multiple projects into shared next-generation sequencing (NGS) runs. We developed a new research workflow to facilitate simultaneous, high-throughput enrichment and NGS of large, multi-locus data sets for two independent phylogenetic studies of the spiny Australian Solanum complex (Solanaceae) and the North American Cordilleran species of Draba (Brassicaceae), respectively. Previous investigations of these taxa have recovered poor phylogenetic resolution with only a few commonly employed loci (e.g., ITS, trnL-trnF, matK-trnK), and additional loci [e.g., single-copy nuclear (SCN) loci] with faster rates of sequence evolution are needed to confidently reconstruct their evolutionary histories. We employed a newly developed tool, MarkerMiner 1.0, and six transcriptomes each from Solanum and Draba from the One Thousand Plants Project (1kp) to identify a set of shared set of SCN loci for both studies. MarkerMiner identified a large set of orthologous SCN genes and, from these, we selected a subset of loci with intronic regions to design and validate primers for high-throughput, targeted enrichment with the Fluidigm Access Array System. Our research workflow—which eliminates redundancy in primer design and validation steps, maximizes sequencing efficiency on NGS platforms, and reduces overall laboratory costs—represents an effective strategy for large-scale data acquisition that can be applied in other laboratories with two or more active phylogenetic studies.
Jordon-Thaden, I., Uribe-Convers, S., Godden, G.T., Srikar, C., Tank, D.C., Martine, C.T. "A Research Workflow for Development of Single-Copy Nuclear Loci and High-throughput Targeted Enrichment for Two or More Phylogenetic Studies." Botany Society of America (2014):