Total RNA was isolated from mouse testes using the mirVana™ miRNA Isolation Kit (Life Technologies). Total RNA samples were treated with Turbo DNase (Invitrogen) and RNA cleanup was done following the manufacturer's instructions using the RNeasy Mini Kit (Qiagen). Strand-specific RNA sequencing libraries were prepared as previously described (Zhang et al., 2012). In brief, total RNA was isolated from frozen koala or mouse tissue samples via the mirVana miRNA Isolation kit and then rRNA depleted using the RiboZero™ Gold rRNA Removal Kit for human, mouse, and rat (Illumina). Mouse total RNA samples were rRNA depleted as previously described (Adiconis et al., 2013; Morlan et al., 2012).