ZFIN Image: Zaksauskaite et al., 2021, Fig. 2

Image

Figure Caption

Fig. 2 Generation and validation of <italic>tdp1<sup>−/−</sup></italic> zebrafish using the CRISPR-Cas9 system.

(A) Sequence of oligonucleotide used for guide RNA (gRNA) synthesis. The scaffolding sequence is in purple; the target sequence is in green, and the T7 polymerase promoter is in blue. (B) Intron-exon structure of the D. rerio tdp1 gene. Exon 2 (in red) was targeted for mutation by Cas9; scale bar, 5000 bp. Photo credit: Ringaile Zaksauskaite, University of Sheffield (generated using http://wormweb.org/exonintron). (C) Sequences of the target region in tdp1^WT zebrafish and two isolated deletion alleles, tdp1^SH475 and tdp1^SH476. The 5-bp deletion (SH476; light blue box) and the 4-bp deletion (SH475; green square). (D) Tdp1^−/+ zebrafish were crossed and genotyped at adulthood; χ² = 2.941 with two degrees of freedom; two-tailed P value of 0.5316. (E) Diagram depicting the TDP1 activity assay showing a 5′ labeled oligomer with a 3′-phosphotyrosyl (PY) that is incubated with zebrafish protein lysate. Active TDP1 processes the 3′-PY into a phosphate group, resulting in a band shift on a DNA sequencing gel. (F) TDP1 activity assay was performed on 600 ng of lysate from 4-dpf embryos. (G) TDP1 activity assay was performed on fin clips from adult zebrafish. LB, lysis buffer control

Figure Data

Phenotype 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 @ Sci Adv