Complete chloroplast genome of Macadamia integrifolia confirms the position of the Gondwanan early-diverging eudicot family Proteaceae

Abstract

Background: Sequence data from the chloroplast genome have played a central role in elucidating the evolutionary history of flowering plants, Angiospermae. In the past decade, the number of complete chloroplast genomes has burgeoned, leading to well-supported angiosperm phylogenies. However, some relationships, particulary among early-diverging lineages, remain unresolved. The diverse Southern Hemisphere plant family Proteaceae arose on the ancient supercontinent Gondwana early in angiosperm history and is a model group for adaptive radiation in response to changing climatic conditions. Genomic resources for the family are limited, and until now it is one of the few early-diverging 'basal eudicot' lineages not represented in chloroplast phylogenomic analyses.

Results: The chloroplast genome of the Australian nut crop tree Macadamia integrifolia was assembled de novo from Illumina paired-end sequence reads. Three contigs, corresponding to a collapsed inverted repeat, a large and a small single copy region were identified, and used for genome reconstruction. The complete genome is 159,714 bp in length and was assembled at deep coverage (3.29 million reads; ~2000 x). Phylogenetic analyses based on 83-gene and inverted repeat region alignments, the largest sequence-rich datasets to include the basal eudicot family Proteaceae, provide strong support for a Proteales clade that includes Macadamia, Platanus and Nelumbo. Genome structure and content followed the ancestral angiosperm pattern and were highly conserved in the Proteales, whilst size differences were largely explained by the relative contraction of the single copy regions and expansion of the inverted repeats in Macadamia.

Conclusions: The Macadamia chloroplast genome presented here is the first in the Proteaceae, and confirms the placement of this family with the morphologically divergent Plantanaceae (plane tree family) and Nelumbonaceae (sacred lotus family) in the basal eudicot order Proteales. It provides a high-quality reference genome for future evolutionary studies and will be of benefit for taxon-rich phylogenomic analyses aimed at resolving relationships among early-diverging angiosperms, and more broadly across the plant tree of life.

Figure 1

Chloroplast genome map of Macadamia integrifolia. Organisation of the chloroplast genome of Macadamia integrifolia (Proteaceae) showing annotated genes. The inner circle shows the location of the large and small single copy (LSC and SSC) and inverted repeat regions (IR_A and IR_B).

Figure 2

Sequence identity plot comparing the chloroplast genome of Macadamia to other Proteales. Pairwise comparisons between Macadamia integrifolia and chloroplast genomes from the Proteales genera Platanus (top) and Nelumbo (bottom) using mVISTA. The y-axis represents % identity ranging from 50-100%. Coding and non-coding regions are marked in purple and pink respectively. Also shown are the genes discussed in the text (red), large single copy (LSC, green), small single copy (SSC, blue) and inverted repeat (IR, grey) regions.

Figure 3

Chloroplast gene Angiospermae phylogeny. Phylogram of the best tree determined by RAxML for the 83-gene, 87-taxa, 49-partition data set. Numbers associated with branches are maximum likelihood percentage bootstrap support values for partitioned and unpartitioned (in italics) analyses. Unnumbered branches had 100% support. Collapsed monophyletic clades and number of taxa in brackets are monocots (11), superasterids (26), superosids (27), magnoliids (3) and gymnosperm outgroup (3). Scale represents substitutions per site.

Figure 4

Inverted repeat Eudicotyledoneae phylogeny. Phylogram of the best tree determined by RAxML for the inverted repeat, 160-taxa, 5-partition data set. Numbers associated with branches are maximum likelihood percentage bootstrap support values for partitioned and unpartitioned (in italics) analyses. Unnumbered branches had 100% support. Collapsed monophyletic clades and number of taxa in brackets are superasterids (59), superosids (82) and Dilleniaceae (7). Scale represents substitutions per site.