A New Approach Using Targeted Sequence Capture for Phylogenomic Studies across Cactaceae

Abstract

Relationships within the major clades of Cactaceae are relatively well known based on DNA sequence data mostly from the chloroplast genome. Nevertheless, some nodes along the backbone of the phylogeny, and especially generic and species-level relationships, remain poorly resolved and are in need of more informative genetic markers. In this study, we propose a new approach to solve the relationships within Cactaceae, applying a targeted sequence capture pipeline. We designed a custom probe set for Cactaceae using MarkerMiner and complemented it with the Angiosperms353 probe set. We then tested both probe sets against 36 different transcriptomes using Hybpiper preferentially retaining phylogenetically informative loci and reconstructed the relationships using RAxML-NG and Astral. Finally, we tested each probe set through sequencing 96 accessions, representing 88 species across Cactaceae. Our preliminary analyses recovered a well-supported phylogeny across Cactaceae with a near identical topology among major clade relationships as that recovered with plastome data. As expected, however, we found incongruences in relationships when comparing our nuclear probe set results to plastome datasets, especially at the generic level. Our results reveal great potential for the combination of Cactaceae-specific and Angiosperm353 probe set application to improve phylogenetic resolution for Cactaceae and for other studies.

Keywords: Hyb-seq; Opuntia; cacti; neotropics; target enrichment.

Figure 1

Flowchart of the pipeline with general steps and tools. For more details see the Methods Section.

Figure 2

Heatmap plot showing recovery efficiency. Columns represent the targeted loci grouped based on probe sets and rows represent the 36 transcriptomes grouped based on the main groups in Cactaceae (OG: outgroup). (a) Length proportion of the target sequences recovered. (b) Size of the target gene sequences recovered.

Figure 3

RAxML best-scoring ML tree of the concatenated database of the two probe sets. All nodes have ≥95% support value bootstrap unless noted. OG: Outgroups; Teph.: Tephrocacteae; C. Cact.: Core Cactoideae.

Figure 4

ASTRAL species tree with pie charts on the nodes representing gene tree topology proportions: blue represents concordance with the shown topology, green is the top alternative bipartition; red is all the other alternative bipartitions; and grey is no support for conflicting bipartition. Branch values represent concordance (top) and conflicting (bottom) gene trees. Asterisks mark all nodes with a posterior probability of ≤90 and scale bars represent coalescent units. (a) C120 and A353 probe sets combined. OG: Outgroups; Teph.: Tephrocacteae; C. Cact.: Core Cactoideae. (b) C120 probe set. (c) A353 probe set.