Embryonic Transcriptome Analysis of E. rowelli

Library preparation and assembly of the embryonic transcriptomes from E. rowelli were performed as described previously [58] (link). Local tBLASTn searches [73] (link) were conducted using transcriptome libraries from different embryonic stages [58] (link). Previously published sequences from other onychophoran and arthropod species were used as queries [29] (link), [74] (link), [75] (link). RNA was isolated from pooled embryos of different developmental stages using TRIzol Reagent (Invitrogen) and RNeasy MinElute Cleanup Kit (Qiagen, Hilden, Germany) according to the manufacturers’ protocols. First-strand synthesis was performed using random hexamer primers and Superscript III polymerase (Invitrogen). Second-strand synthesis was carried out with DNA Pol I polymerase (Invitrogen). The obtained cDNA was purified using NucleoSpin Extract II-Kit (Macherey-Nagel, Düren, Germany) following the manufacturer's protocol. Fragments of engrailed, cubitus interruptus, wingless and hedgehog were amplified using specific primers (Table 1). The corresponding sequences were made available under the GenBank accession numbers KF218600–KF218603.