DOI: 10.5061/dryad.bk3j9kdc0
title: Data from: Opsins in Onychophora (velvet worms) suggest a single origin and subsequent diversification of visual pigments in arthropods

description: Multiple visual pigments, prerequisites for color vision, are found in
 arthropods, but the evolutionary origin of their diversity remains
 obscure. In this study, we explore the opsin genes in five distantly
 related species of Onychophora, using deep transcriptome sequencing and
 screening approaches. Surprisingly, our data reveal the presence of only
 one opsin gene (onychopsin) in each onychophoran species, and our
 behavioral experiments indicate a maximum sensitivity of onychopsin to
 blue–green light. In our phylogenetic analyses, the onychopsins represent
 the sister group to the monophyletic clade of visual r-opsins of
 arthropods. These results concur with phylogenomic support for the
 sister-group status of the Onychophora and Arthropoda and provide evidence
 for monochromatic vision in velvet worms and in the last common ancestor
 of Onychophora and Arthropoda. We conclude that the diversification of
 visual pigments and color vision evolved in arthropods, along with the
 evolution of compound eyes—one of the most sophisticated visual systems
 known.
description: Total RNA of five onychophoran species, including three species
 of Peripatopsidae (<em>Euperipatoides rowelli</em>,
 <em>Phallocephale tallagandensis</em>, and
 <em>Ooperipatus hispidus</em>) and two species of Peripatidae
 (<em>Principapillatus hitoyensis</em> [referred to
 as <em>Epiperipatus</em> cf. <em>isthmicola</em>
 in the published article], and <em>Eoperipatus</em> sp.) was
 extracted using TRIzol (Invitrogen, Carlsbad, CA) and the RNeasy MinElute
 Cleanup Kit (Qiagen, Hilden, Germany) according to the manufacturers
 protocols. Double-stranded complementary DNA was prepared following
 Illuminas messenger RNA (mRNA)-seq sample preparation guide (Part #1004898
 Rev. D). Briefly, mRNA was isolated from total RNA using Dynal oligo(dT)
 beads (Invitrogen) and fragmented (~200bp) using divalent cationic ions.
 First-strand synthesis was performed with random hexamer primers, using
 Superscript II polymerase (Invitrogen). Subsequent second-strand synthesis
 was carried out with DNA Pol I polymerase (Invitrogen). Starting at the
 blunt end repair step, multiplex sequencing libraries were prepared
 following the multiplex protocol of Meyer and Kircher (2010), with
 modifications for double indexing described in Kircher et al. (2012).
 These libraries were sequenced according to the manufacturers instructions
 with 75 cycles paired end on the Genome Analyzer IIx platform (v4
 sequencing chemistry and v4 cluster generation kit; Illumina, San Diego,
 CA). Raw sequences were analyzed with IBIS 1.1.2
 (Kircher et al. 2009). For highly accurate sample identification,
 sequences with falsely paired indices were discarded. Paired-end reads
 from a single cluster were merged, if at least 11 bp were overlapping
 (Kircher et al. 2011). Reads with low complexity were removed. For each
 library from the five species, we trimmed the Illumina raw data at three
 different levels of stringency (Filter15: reads with more than five bases
 below a phred quality score of 15 were removed; Filter30: reads with less
 than 20 bases in a row with a phred quality score of at least 30 were
 removed; Filter25: same as Filter30 but the threshold value was set to 25
 to keep as many high-quality reads as possible, 63–89% compared with
 Filter15). This was done to deal with the tradeoff between the quality and
 quantity of reads as we lost many reads (51–90% compared with Filter15) by
 using the very stringent Filter30, which thus might have led to a loss of
 lowly expressed genes (for details see statistics_and_BUSCO_values.xls).
 For all libraries and filters described, sequence reads were assembled
 <em>de novo</em> using the CLC Genomics Workbench 5.1 (CLC
 bio, Århus, Denmark) with the following parameters: mismatch cost 3;
 insertion cost 3; deletion cost 3; length fraction 0.5; similarity
 fraction 0.8; minimum contig length 200; automatic word size; automatic
 bubble size; and contig adjustment by mapped reads (for details see
 statistics_and_BUSCO_values.xls).
description: An explanation of each file is given in the README.txt file.