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Development of single-copy nuclear intron markers for species-level phylogenetics: Case study with Paullinieae (Sapindaceae).

ABSTRACT: PREMISE OF THE STUDY:We developed a bioinformatic pipeline that leverages a publicly available genome and published transcriptomes to design primers in conserved coding sequences flanking targeted introns of single-copy nuclear loci. Paullinieae (Sapindaceae) is used to demonstrate the pipeline. METHODS AND RESULTS:Transcriptome reads phylogenetically closer to the lineage of interest are aligned to the closest genome. Single-nucleotide polymorphisms are called, generating a "pseudoreference" closer to the lineage of interest. Several filters are applied to meet the criteria of single-copy nuclear loci with introns of a desired size. Primers are designed in conserved coding sequences flanking introns. Using this pipeline, we developed nine single-copy nuclear intron markers for Paullinieae. CONCLUSIONS:This pipeline is highly flexible and can be used for any group with available genomic and transcriptomic resources. This pipeline led to the development of nine variable markers for phylogenetic study without generating sequence data de novo.

SUBMITTER: Chery JG

PROVIDER: S-EPMC5628028 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Development of single-copy nuclear intron markers for species-level phylogenetics: Case study with Paullinieae (Sapindaceae).

Chery Joyce G JG Sass Chodon C Specht Chelsea D CD

Applications in plant sciences 20170928 9

<h4>Premise of the study</h4>We developed a bioinformatic pipeline that leverages a publicly available genome and published transcriptomes to design primers in conserved coding sequences flanking targeted introns of single-copy nuclear loci. Paullinieae (Sapindaceae) is used to demonstrate the pipeline.<h4>Methods and results</h4>Transcriptome reads phylogenetically closer to the lineage of interest are aligned to the closest genome. Single-nucleotide polymorphisms are called, generating a "pseu ...[more]

PMID: 28989824

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Project description:Background and aimsThe tribe Paullinieae has the highest diversity of vascular variants among the seed plants. The developmental diversity is better understood in the species-rich genera Paullinia and Serjania; however, the phylogeny and diversity of vascular variants in the smaller genera of Paullinieae remain understudied. Here we investigate the evolution of development of stem vasculatures in the small genus Urvillea.MethodsWe generate the first molecular phylogeny of Urvillea derived from 11 markers using a maximum likelihood and Bayesian approach. In combination with phylogenetic reconstruction, stochastic character mapping is used to assess evolutionary changes in stem ontogenies, determined from developmental anatomy of stems collected in the field or from herbarium and wood collections.Key resultsUrvillea is supported as a monophyletic group and sister to Serjania. There are five stem ontogenies in Urvillea, including typical growth and four different vascular variants. Most stem ontogenies initiate with lobed stems in primary growth. Lobed stems in secondary growth are ancestral in Urvillea, but this ontogeny was lost multiple times. A reversal to typical growth occurred in non-climbing species. Phloem wedges, fissured stems, and ectopic cambia each evolved once independently. Phloem wedges is an intermediate developmental stage in the formation of fissured stems, which is characterized by a continuous fragmentation of vascular tissues. Lobed stems may generate constriction zones and lobes may split or not.ConclusionsUrvillea is the third most diverse genus (after Serjania and Paullinia) with respect to the number of vascular variants within Paullinieae. One ontogeny (fissured stems) is exclusive to the genus. Differential cambial activity and ectopic cambia are the main ontogenetic processes generating stem diversity. The evolutionary history of vascular variants demonstrates the large developmental plasticity of the cambium in such a small genus and further demonstrates that complex anatomies have repeatedly evolved within Paullinieae lianas.

Project description:Nucleotide sequences representing nine genes and five presumptive genetic loci were used to infer phylogenetic relationships among seven Baylisascaris species, including one species with no previously available molecular data. These genes were used to test the species status of B. procyonis and B. columnaris using a coalescent approach. Phylogenetic analysis based on combined analysis of sequence data strongly supported monophyly of the genus and separated the species into two main clades. Clade 1 included B. procyonis, B. columnaris, and B. devosi, species hosted by musteloid carnivores. Clade 2 included B. transfuga and B. schroederi from ursids, B. ailuri, a species from the red panda (a musteloid), and B. tasmaniensis from a marsupial. Within clade 2, geographic isolates of B. transfuga, B. schroederi (from giant panda), and B. ailuri formed a strongly supported clade. In certain analyses (e.g., some single genes), B. tasmaniensis was sister to all other Baylisascaris species rather than sister to the species from ursids and red panda. Using one combination of priors corresponding to moderate population size and shallow genetic divergence, the multispecies coalescent analysis of B. procyonis and B. columnaris yielded moderate support (posterior probability 0.91) for these taxa as separate species. However, other prior combinations yielded weak or no support for delimiting these taxa as separate species. Similarly, tree topologies constrained to represent reciprocal monophyly of B. columnaris and B. procyonis individuals (topologies consistent with separate species) were significantly worse in some cases, but not others, depending on the dataset analyzed. An expanded analysis of SNPs and other genetic markers that were previously suggested to distinguish between individuals of B. procyonis and B. columnaris was made by characterization of additional individual nematodes. The results suggest that many of these SNPs do not represent fixed differences between nematodes derived from raccoon and skunk hosts.

Project description:BackgroundMolecular phylogenetic investigations have revolutionized our understanding of the evolutionary history of ferns-the second-most species-rich major group of vascular plants, and the sister clade to seed plants. The general absence of genomic resources available for this important group of plants, however, has resulted in the strong dependence of these studies on plastid data; nuclear or mitochondrial data have been rarely used. In this study, we utilize transcriptome data to design primers for nuclear markers for use in studies of fern evolutionary biology, and demonstrate the utility of these markers across the largest order of ferns, the Polypodiales.Principal findingsWe present 20 novel single-copy nuclear regions, across 10 distinct protein-coding genes: ApPEFP_C, cryptochrome 2, cryptochrome 4, DET1, gapCpSh, IBR3, pgiC, SQD1, TPLATE, and transducin. These loci, individually and in combination, show strong resolving power across the Polypodiales phylogeny, and are readily amplified and sequenced from our genomic DNA test set (from 15 diploid Polypodiales species). For each region, we also present transcriptome alignments of the focal locus and related paralogs-curated broadly across ferns-that will allow researchers to develop their own primer sets for fern taxa outside of the Polypodiales. Analyses of sequence data generated from our genomic DNA test set reveal strong effects of partitioning schemes on support levels and, to a much lesser extent, on topology. A model partitioned by codon position is strongly favored, and analyses of the combined data yield a Polypodiales phylogeny that is well-supported and consistent with earlier studies of this group.ConclusionsThe 20 single-copy regions presented here more than triple the single-copy nuclear regions available for use in ferns. They provide a much-needed opportunity to assess plastid-derived hypotheses of relationships within the ferns, and increase our capacity to explore aspects of fern evolution previously unavailable to scientific investigation.

Dataset Information

Development of single-copy nuclear intron markers for species-level phylogenetics: Case study with Paullinieae (Sapindaceae).

Publications

Development of single-copy nuclear intron markers for species-level phylogenetics: Case study with Paullinieae (Sapindaceae).

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