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Abstract Detail


Folk, Ryan [1], Mandel, Jennifer [2], Freudenstein, John [3].

Protocol for Targeted Enrichment of Intron-Containing Sequence Markers for Recent Radiations: A Phylogenomic Example from Heuchera (Saxifragaceae) .

Phylogenetic inference is moving to increasingly large multilocus datasets that incorporate information from numerous nuclear and organellar loci. The performance of Next-Generation Sequencing methods at the boundary between population and phylogenetic relationships requires methodological refinement and case studies. To address this gap, we present a method for targeting long loci spanning intron-exon boundaries, using Heuchera as a test case where Sanger-based markers failed to obtain support for many relationships. 278 loci (average length 1362 bp) were designed using a splice-site prediction method with transcriptomic and genomic data from two species of Heuchera. Additionally complete mitochondrial and chloroplast genomes were assembled from the genomic data for processing off-target reads. Biotinylated probes were designed for a target enrichment experiment with 15 multiplexed individuals on an Illumina MiSeq lane. Sequence assembly was performed with a reference-based method, using the contigs from the locus design step as genomic references; additionally chloroplast and mitochondrial data used as references for off-target reads. Complete or nearly complete (> 99%) sequences were assembled from essentially all loci from all taxa, with a final matrix that was ~2/3 intronic. Aligned introns showed a four-fold increase in divergence as opposed to exons, and > 40 fold increase in gap content. Concatenated analysis gave decisive support to a preliminary phylogeny, including nodes that were recalcitrant with Sanger-based markers, and support was also high and relationships mostly similar in the coalescent analysis. The use of off-target reads resolved phylogenetic hypotheses for the organelles, each of which is in strong conflict with the nuclear signal. Our approach shows promise in resolving a recent radiation that was recalcitrant using Sanger-based sequence markers. Intron targeting is highly successful at low taxonomic levels despite higher substitution and indel frequencies, and should be exploited in studies of species complexes.

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1 - The Ohio State University, 1315 Kinnear Road, Columbus, OH, 43220, USA
2 - University Of Memphis, 3744 Walker Ave, Memphis, TN, 38152, USA
3 - Ohio State University, Museum Of Biological Diversity, 1315 Kinnear Rd, Columbus, OH, 43212-1157, USA

targeted enrichment
probe capture
Rapid radiation
marker development.

Presentation Type: Oral Paper:Papers for Topics
Session: 32
Location: Salon 11/The Shaw Conference Centre
Date: Tuesday, July 28th, 2015
Time: 8:30 AM
Number: 32003
Abstract ID:259
Candidate for Awards:None

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