Supplementary data for: Transcriptomics of mosaic brain differentiation underlying complex division of labor in a social insect

Concerted developmental programming may constrain changes in component structures of the brain, thus limiting the ability of selection acting on individual brain compartments to form an adaptive mosaic independent of total brain size or body size. Measuring patterns of gene expression underpinning brain scaling in conjunction with anatomical brain atlases can aid in identifying influences of concerted and/or mosaic evolution. Species exhibiting exceptional size and behavioral polyphenisms provide excellent systems to test predictions of brain evolution models by quantifying brain gene expression. We examined patterns of brain gene expression in a remarkably polymorphic and behaviorally complex social insect, the leafcutter ant Atta cephalotes. Approximately ~50% of differential gene expression observed among three morphologically, behaviorally, and neuroanatomically differentiated worker size groups was attributable to body size, but we also found strong evidence of differential brain g..., Brains of mature, fully sclerotized Atta cephalotes workers categorized by size group as minims (0.5–0.7mm in head width), medias (1.7–1.9mm in head width), and majors (≥3mm in head width) were sampled from three mature colonies for gene expression analyses. We prepared between nine and 11 samples from each worker size group, composed of three to 10 pooled brains depending on worker size group, distributed across three colonies of origin, for a total of 30 samples.
Total RNA was extracted from worker brains using a ThermoFisher PicoPure kit. Sample quality and quantity, as well as lack of protein or DNA contaminants, were assessed using a Thermo Scientific Nanodrop spectrophotometer and an Agilent Bioanalyzer 2100, respectively.
Libraries were sequenced using a combination of Illumina NextSeq and MiSeq with SE 75 reads. RNAseq unstranded libraries with mRNA poly-A selection were prepared by Harvard BioPolymers using a KAPA mRNA HyperPrep kit. mRNA sequence libraries were individually ba...,