Abstract
BackgroundMolecular evolutionary studies share the common goal of elucidating historical relationships, and the common challenge of adequately sampling taxa and characters. Particularly at low taxonomic levels, recent divergence, rapid radiations, and conservative genome evolution yield limited sequence variation, and dense taxon sampling is often desirable. Recent advances in massively parallel sequencing make it possible to rapidly obtain large amounts of sequence data, and multiplexing makes extensive sampling of megabase sequences feasible. Is it possible to efficiently apply massively parallel sequencing to increase phylogenetic resolution at low taxonomic levels?ResultsWe reconstruct the infrageneric phylogeny of Pinus from 37 nearly-complete chloroplast genomes (average 109 kilobases each of an approximately 120 kilobase genome) generated using multiplexed massively parallel sequencing. 30/33 ingroup nodes resolved with ≥ 95% bootstrap support; this is a substantial improvement relative to prior studies, and shows massively parallel sequencing-based strategies can produce sufficient high quality sequence to reach support levels originally proposed for the phylogenetic bootstrap. Resampling simulations show that at least the entire plastome is necessary to fully resolve Pinus, particularly in rapidly radiating clades. Meta-analysis of 99 published infrageneric phylogenies shows that whole plastome analysis should provide similar gains across a range of plant genera. A disproportionate amount of phylogenetic information resides in two loci (ycf1, ycf2), highlighting their unusual evolutionary properties.ConclusionPlastome sequencing is now an efficient option for increasing phylogenetic resolution at lower taxonomic levels in plant phylogenetic and population genetic analyses. With continuing improvements in sequencing capacity, the strategies herein should revolutionize efforts requiring dense taxon and character sampling, such as phylogeographic analyses and species-level DNA barcoding.
Highlights
Molecular evolutionary studies share the common goal of elucidating historical relationships, and the common challenge of adequately sampling taxa and characters
Molecular phylogenetic and phylogeographic analyses are typically limited by DNA sequencing costs, and this forces investigators to choose between dense taxon sampling with a small number of maximally informative loci, or genome-scale sampling across a sparse taxon sample [14]
In this paper we demonstrate the feasibility and effectiveness of Massively parallel sequencing (MPS)-based chloroplast phylogenomics for onethird of the world's pine species (Pinus), a lineage with numerous unresolved relationships based on previous chloroplast DNA (cpDNA)-based studies [20,21,22]
Summary
Molecular evolutionary studies share the common goal of elucidating historical relationships, and the common challenge of adequately sampling taxa and characters. Molecular phylogenetic and phylogeographic analyses are typically limited by DNA sequencing costs, and this forces investigators to choose between dense taxon sampling with a small number of maximally informative loci, or genome-scale sampling across a sparse taxon sample [14] Balancing these choices is difficult in studies focused on recently diverged taxa or ancient rapid radiations, as taxon sampling needs to be sufficiently large to define the magnitude of intraspecific variation and the SDJHQXPEHUQRWIRUFLWDWLRQSXUSRVHV. These barriers have been largely eliminated through four innovations: development of strategies for targeted isolation of large genomic regions [9,13,14,15]; harnessing the capacity of these platforms to sequence targeted regions in multiplex [9,14,16]; streamlining sample preparation and improving throughput [17]; and developing accurate de novo assemblers that reduce reliance upon a predefined reference sequence [18,19]
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