10.25386/genetics.7624559.v1
Amhed M. Vargas Velazquez
Amhed M. Vargas
Velazquez
Fabrice Besnard
Fabrice
Besnard
Marie-Anne Félix
Marie-Anne
Félix
Supplemental Material for Vargas-Velazquez, Besnard, and Félix, 2019
GSA Journals
2019
Oscheius tipulae
vulva development
evolution
nematode
genetic screen
Genetics
2019-01-30 16:27:14
Dataset
https://gsajournals.figshare.com/articles/dataset/Supplemental_Material_for_Vargas-Velazquez_Besnard_and_F_lix_2019/7624559
<p><b>Figure S1. Dissecting microscope pictures of wild-type and
mutant <i>O. tipulae</i> adult hermaphrodites.</b></p>
<p>WT: wild-type. <i>Oti-lin-3(mf86)</i>: fully-penetrant egg-laying
defective, forming a bag of worms. <i>Oti-mom-5(sy493)</i>: also partially egg-laying
defective and protruding vulva. <i>Oti-plx-1(mf78</i>): protruding vulva. All the
images are set to the same scale. Scale bar: 100 micrometers.</p>
<p> </p>
<p><b>Figure S2. Example of mapping-by-sequencing in <i>O. tipulae.</i></b></p>
<p>Graphs showing the frequency of JU170 calls for
single-nucleotide polymorphisms between CEW1 (reference wild isolate, on which
the mutagenesis was conducted) and JU170 (alternative wild isolate used for
mapping) along scaffold 10 of genome assembly nOt.2.0, for two alleles of the
<i>cov-4</i> locus, called <i>sy465</i> and <i>sy493</i>. The location of <i>Oti-mom-5 </i>is marked by a
grey line. See Besnard et al. 2017 for further details.</p>
<p> </p>
<p><b>Figure S3. Single-molecule FISH of Wnt genes</b>.</p>
<p>A) Examples of the localization of WNT mRNAs as revealed by
fluorescence and DAPI signal. The size of the bar is 10 micrometers. B) Average
number of mRNAs of WNT genes in the L3 stage along an antero-posterior axis
defined from the beginning of the gut to the last nucleus seen in the tail by
DAPI.</p>
<p> </p>
<p><b>Figure S4. Phylogenetic relationship between Wnt genes
inside and outside the Caenorhabditis clade.</b></p>
<p>The cladograms were inferred using the Neighbor-Joining
method with 1000 replicates for boostrapping. The percentage of replicate trees
in which the associated taxa clustered together in the bootstrap test are shown
next to the branches. Evolutionary analyses were conducted in MEGA X.
Abbreviations: <i>Cbr (C. briggsae), Cel (C. elegans), Cjp (C. japonica), Dmel (Drosophila
melagonaster), Oti (O. tipulae), Ovo (Onchocerca volvulus)</i>.</p>
<p> </p>
<p><b>Figure S5. Identification and phylogenetic relationships of
Delta/Serrate/Lag-2 (DSL) proteins in <i>O. tipulae</i>.</b></p>
<p>A) Cladogram inferred using the Neighbor-Joining method with
1000 replicates for boostrapping. Abbreviations: <i>Cbr (C. briggsae), Cel (C.
elegans), Cjp (C. japonica), Dm (Drosophila melagonaster), Oti (O. tipulae),
Ppa (Pristionchus pacificus)</i>.</p>
<p>B) Alignment of the delta motif used to calculate the
molecular distances between DSL proteins.</p>
<p> </p>
<p><b>Figure S6. Expression profile of <i>Cel-lin-44</i> revealed by
smFISH.</b></p>
<p>A) Expression of <i>lin-44</i> (red) in the sex myoblast precursors
(identified with the <i>hlh-8::GF</i>P transgene, in blue) before P6.p division.</p>
<p>B) Expression of <i>lin-44</i> (red) in the sex myoblast precursors
(blue) and in P6.p daughters. The anchor cell is labeled with probes against
<i>lag-2</i> (green).</p>
<p>The scale bar measures 10 micrometers and is the same for
both images.</p>
<p> </p>
<p><b>Figure S7. Cis-regulation of <i>Cel-lin-3</i> and <i>Oti-lin-3
</i>expression.</b></p>
<p>A) Diagram representing binding sites (rectangles) known to
be recurrent in the upstream region of <i>Caenorhabditis</i> species <i>lin-3</i> (Barkoulas
et al. 2016). The orientation of the binding sequence is defined by its
position with respect to the line (i.e., above for the forward strand and below
for the reverse strand).</p>
<p>B) <i>Oti-lin-3</i> expression patterns revealed by smFISH in
wild-type and <i>Oti-lin-3(mf86)</i> animals. The animals are shown in ventral view,
thus showing the developing gonad primordium on either side of the anchor
cell. The arrows indicate the position
of the anchor cell as revealed by the presence of<i> Oti-dsl</i> mRNA. The scale bar
measures 10 micrometers and is the same for both images.</p>
<p> </p>
<p><b>Table S1. List of strains used in this study.</b></p><b>
</b><p><b>Table S2. Sequences of DNA primers used in this study.</b></p>
<p>Sequencing primers to verify by Sanger sequencing the
mutations identified by the mapping by sequencing approach, and to identify the
molecular lesion in additional alleles.</p>
<p><b>Table S3. Sequences of smFISH probes used in this study.</b></p>
<p>The fluorophore coupled to each probe is noted at the end of
the set name.</p>
<p><b>Table S4. smFISH quantifications, distance measurements and
vulval cell fates used in this study.</b></p>
<p> </p>