Phylogeographic patterns of the Australian grass trees (

Abstract

Despite the prevalence of sclerophyllous vegetation in Australia, there are few phylogeographic studies with continent-wide sampling addressing questions about the expansion of this vegetation post-aridification, and what physical barriers are associated with breaks in distribution or patterns of genetic structure. We used amplicon sequencing of plastid and nuclear DNA to investigate phylogeography of Xanthorrhoea. Despite extremely low genetic variation and widespread sharing of identical DNA sequence among multiple species and across significant geographic distance, haplotype analyses showed phylogeographic structure. Network analysis of six plastid loci and 137 samples identified 54 haplotypes (two common, 35 unique to single samples) in three geographic groups, south west and central Australia, northern–eastern Australia, and southern–eastern Australia, indicating restricted gene flow among regions. A nrDNA haplotype network of 152 samples showed less variation, with one haplotype being widespread, found in all three plastid geographic groups and 70% of species. nrDNA indicated four groups in a pattern, suggestive of at least two east–west divergences. A geographic area of a high haplotype diversity and divergence was identified in southern New South Wales, near the Southern Transition Zone. Our results showed the value of targeted amplicon sequencing and using multiple samples per species to identify both broad-scale and fine-scale biogeographic patterns.

Phylogeny and classification of the Australasian and Indomalayan mimosoid legumes Archidendron and Archidendropsis (Leguminosae, subfamily Caesalpinioideae, mimosoid clade)

“…Sequences were aligned in Geneious v.8.1.4 (Biomatters Ltd.) and assessed for variability between the samples. The most variable loci were then used in a targeted amplicon sequencing ( TAS ) approach ( McLay et al 2021 ), sequencing pooled amplicons on an Illumina MiSeq. For this, additional internal primers were designed for the five loci that had a total amplicon length greater than 500 bp, in order to produce shorter amplicons that could be fully sequenced using a 500-cycle sequencing kit.…”

“…Library preparation followed the two-step PCR process outlined in McLay et al (2021) . The first step used the region-specific primers to amplify each locus individually for each sample.…”

Species-specific basecallers improve actual accuracy of nanopore sequencing in plants

“…We sought to assess if basecalling could be improved by using a single-species and genome-specific trained basecalling model by training basecalling models using R9.4.1 flowcells for both Phebalium stellatum (eudicot, Sapindales, Rutaceae; [ 16 ]) and Xanthorrhoea johnsonii (monocot, Asparagales, Asphodelaceae; [ 17 ]), and R10.4 flowcells for P. stellatum, two Australian plants whose lineages diverged ~ 136 million years ago [ 18 ]. Additionally, by reciprocally basecalling our two species’ ONT sequencing data ( P. stellatum was basecalled with X. johnsonii’s model and X. johnsonii was basecalled with P. stellatum’s model), we sought to test whether the mixed nature of ONT’s basecalling models could be affecting basecalling accuracy.…”

Plant species-specific basecaller improves actual accuracy of nanopore sequencing

“…We sought to assess if basecalling could be improved by using a single-species and genome-speci c trained basecalling model by training basecalling models using R9.4.1 owcells for both Phebalium stellatum (eudicot, Sapindales, Rutaceae; [16]) and Xanthorrhoea johnsonii (monocot, Asparagales, Asphodelaceae; [17]), and R10.4 owcells for P. stellatum, two Australian plants whose lineages diverged ~ 136 million years ago [18]. Additionally, by reciprocally basecalling our two species' ONT sequencing data (P. stellatum was basecalled with X. johnsonii's model and X. johnsonii was basecalled with P. stellatum's model), we sought to test whether the mixed nature of ONT's basecalling models could be affecting basecalling accuracy.…”