IMAGE

Figure 7

ID

ZDB-IMAGE-201107-26

Genes

rab13

Publication

Costa et al., 2020 - RAB13 mRNA compartmentalisation spatially orients tissue morphogenesis

All Figures

Figures for Costa et al., 2020

Image

Figure Caption

Figure 7 CRISPR‐Cas9 editing of the zebrafish <italic>rab13</italic> 3′UTR perturbs mRNA polarisation

CRISPR‐Cas9 strategy to generate the Tg(kdrl:EGFP) rab13^+/∆3′UTR zebrafish strain. The wild‐type (Wt) rab13 exon 8 is represented with its coding sequence in dark and the 3′UTR in clear boxes; the 5′ and 3′ gRNA‐targeted regions are represented with green lines. Arrows: relative positions of the forward (f) and reverse (r) PCR primers used to identify animals with CRISPR‐Cas9-mediated deletions (∆) in the rab13 3′UTR.

Representative genotyping PCR demonstrates the band size shift in zebrafish harbouring a ∆482 rab13 3′UTR. Asterisk marks a heteroduplex formed between Wt and ∆482 rab13 3′UTR PCR amplicons.

Detailed DNA sequence depicting nucleotide positions within the Wt and ∆482 rab13 3′UTR.

RNAseq mapped reads depicting rab13 exon usage in Tg(kdrl:EGFP) rab13^+/+ and rab13^{∆3′UTR/∆3′UTR} zebrafish embryos. Coloured lines indicate SNPs.

qPCR analysis of rab13 mRNA levels in individual 26–28 hpf clutch‐matched sibling embryos (n ≥ 9 embryos; ns: not significant; Kruskal–Wallis test with Dunn's correction).

smFISH detection of rab13 and kdr mRNA in cultured GFP‐expressing endothelial cells extracted from 48 hpf Tg(kdrl:EGFP) rab13^+/+ and rab13^{∆3′UTR/∆3′UTR} zebrafish embryos.

Polarisation Index (PI) of rab13 and kdr detected by smFISH in individual zebrafish cells (n ≥ 8 cells; **P < 0.01, ns: not significant; one‐way ANOVA with Bonferroni's correction).

rab13 PI plotted against respective kdr PI. The slope of the coloured lines represents the average rab13/kdr PI ratio; the dashed grey line represents a 1:1 ratio (n ≥ 8 cells).

Data information: +/+, +/∆ and ∆/∆ represent Tg(kdrl:EGFP) rab13^+/+, rab13^+/∆3′UTR and rab13^{∆3′UTR/∆3′UTR} embryos, respectively (E, G, H). Arrows indicate orientation of RNA localisation; yellow dashed lines outline cell borders; red circles highlight smFISH spots; scale bars = 10 μm (F). Bar charts are presented as means ± s.d.

Figure Data

Expression / Labeling details

Acknowledgments

This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ EMBO J.