Testis-specific RNA-Seq of Calligrapha (Chrysomelidae) as a transcriptomic resource for male-biased gene inquiry in Coleoptera.

We report the architecture of testis transcriptomes of four closely related species of Calligrapha (Chrysomelidae) beetles, which diverged during the last 3 million years. Five cDNA libraries were sequenced using Illumina HiSeq technology, retrieving 102 884-176 514 assembled contigs, of which ~33-45% of these longer than 499 nt were functionally annotated. Annotation and sequence similarity comparisons of these libraries revealed high homogeneity in gene composition and the presence of several functional candidates related to reproduction or reproductive processes (0.72-1.08% of annotated sequences). Stringent sequence similarity analyses of these transcriptomes against empirically demonstrated male-biased genes in Drosophila melanogaster and Tribolium castaneum allowed the identification of 77 homologues in Calligrapha, possible candidates of male-biased expression. Some of these genes - including CG9313, Tektin-A or tomboy40 - were confirmed as orthologs of these male-biased genes using phylogenetic inference and available model insect data, increasing our confidence that they represent functional homologues too. Our transcriptomes are a valuable transcriptomic resource for the analysis of male-biased genes in Calligrapha, which has the added interest of including several female-only species. But it simultaneously represents a landmark for similar studies in Coleoptera, broadening the taxonomic diversity currently represented by the model species T. castaneum, and incipient genomic data in other herbivorous lineages, including weevils, longhorn beetles and leaf beetles.