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, 4, e1584
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Revisiting the Zingiberales: Using Multiplexed Exon Capture to Resolve Ancient and Recent Phylogenetic Splits in a Charismatic Plant Lineage

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Revisiting the Zingiberales: Using Multiplexed Exon Capture to Resolve Ancient and Recent Phylogenetic Splits in a Charismatic Plant Lineage

Chodon Sass et al. PeerJ.

Abstract

The Zingiberales are an iconic order of monocotyledonous plants comprising eight families with distinctive and diverse floral morphologies and representing an important ecological element of tropical and subtropical forests. While the eight families are demonstrated to be monophyletic, phylogenetic relationships among these families remain unresolved. Neither combined morphological and molecular studies nor recent attempts to resolve family relationships using sequence data from whole plastomes has resulted in a well-supported, family-level phylogenetic hypothesis of relationships. Here we approach this challenge by leveraging the complete genome of one member of the order, Musa acuminata, together with transcriptome information from each of the other seven families to design a set of nuclear loci that can be enriched from highly divergent taxa with a single array-based capture of indexed genomic DNA. A total of 494 exons from 418 nuclear genes were captured for 53 ingroup taxa. The entire plastid genome was also captured for the same 53 taxa. Of the total genes captured, 308 nuclear and 68 plastid genes were used for phylogenetic estimation. The concatenated plastid and nuclear dataset supports the position of Musaceae as sister to the remaining seven families. Moreover, the combined dataset recovers known intra- and inter-family phylogenetic relationships with generally high bootstrap support. This is a flexible and cost effective method that gives the broader plant biology community a tool for generating phylogenomic scale sequence data in non-model systems at varying evolutionary depths.

Keywords: Ancient radiation; Array-based capture; Banana; Exon capture; Gingers; HTS; Heliconiaceae; High-throughput sequencing; Musaceae.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1
Figure 1. Schematic diagrams for the bioinformatic work flow.
(A) Work flow to generate family specific bait sequence from transcriptomes and the annotated exons from Musa acuminata and (B) work flow to generate individual sequences for each gene from raw reads independent of de novo assembly. Base changes and SNPs are highlighted and the schematic is represented as in the SAMtools tview format (i.e., reverse reads are represented with commas and lowercase letters). The representation is condensed to show examples of how the reads are transformed but the actual coverage used to call SNPs was at least 20× (see methods).
Figure 2
Figure 2. Capture efficiency across individuals and exons.
(A) Average coverage-depth across all individuals as represented by colored bars placed according to location on Musa acuminata chromosomes. Exons with greater than 200× coverage are represented with black bars. Bars representing exons with less than 200× coverage are colored according to average coverage in the scale shown. Exons with unknown chromosomal location are not shown but information can be found in Table S2. (B) Average coverage over all exons for each individual after removing PCR duplicates and with strict alignment parameters that were used for SNP calling. Average coverage was calculated before and after removing the high coverage exons indicated in 2A. (C) Per individual, the percent of the total sequenced base pairs passing Illumina quality filters that mapped to target regions prior to PCR duplicate removal. Percent of base pairs mapping to chloroplast plastid and nuclear regions are indicated in orange and blue, respectively. Species are grouped by family (Can=Cannaceae, Mar=Marantaceae, Cos=Costaceae, Zin=Zingiberaceae, Str=Strelitziaceae, Low=Lowiaceae, Hel=Heliconiaceae, Mus=Musaceae) and species upon which baits were generated are indicated with a filled circle (nuclear bait) or open circle (plastid bait).
Figure 3
Figure 3. Phylogenetic tree of Zingiberales based on a partitioned ML of concatenated plastid and nuclear sequence.
Bootstrap support values adjacent to branches as MP/ML/ASTRAL. A dot indicates 100% bootstrap support, a dash indicates less than 50% support and an asterisk indicates conflicting topology in the ASTRAL optimal tree. Scale is in expected substitutions per site.

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Grant support

This research was supported by NSF DEB 1257701 Collaborative Research Award to CDS and SYS, and NSF DEB 0816661 Research Opportunity Award Supplement to CDS and CFB. This work used the Vincent J. Coates Genomics Sequencing Laboratory at UC Berkeley, supported by NIH S10 Instrumentation Grants S10RR029668 and S10RR027303. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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