formamide solution  (New England Biolabs)

Journal: Molecular Biology and Evolution

Article Title: Genetic and Developmental Divergence in the Neural Crest Program between Cichlid Fish Species

doi: 10.1093/molbev/msae217

Figure Lengend Snippet: Paralog-specific KO phenotypes indicate functional divergence of sox10a and sox10b in cichlids. a) Melanophore pigmentation defects, particularly decreased abundance, are observed in sox10b but not in sox10a A. calliptera mutants. b) sox10b -CRISPR embryos have significantly reduced melanophore counts at 7 dpf compared with WT and control injections (boxplots showing melanophore count within the dorsal head area outlined in a.; P -values are shown for Tukey HSD). c) Representative craniofacial phenotypes of sox10a- CRISPR embryos in live animals and their corresponding cartilage preparations in lateral and ventral views. White dashed lines show the reduced frontal slope of the brain case. Arrowheads and asterisks in panels showing cartilage preparations indicate reduced and missing cartilages, respectively, in the mutants. Fractions indicate the frequency of presented phenotype across all surviving embryos on that day. Data shown are representative of at least biological replicates per target gene. Ba, branchial arches; bhy, basihyal; dpf, days postfertilization; op, orbital process; pf, pectoral fin; soc, super-orbital cartilages; WT, wild-type. Scale bars = 1 mm.

Article Snippet: Embryos were stained for cartilage following the protocol of with following modifications: (i) all specimens were bleached to remove melanophore pigmentation using a solution of 0.05% hydrogen peroxide (Sigma) and 0.05% formamide (ThermoFisher) for 30 to 45 min under light and (ii) samples were cleared using first 50% then 70% glycerol:water solutions until complete sinking.

Techniques: Functional Assay, CRISPR, Control