Fig 2

Intronic insertion of the T3H48 ribozyme in the alb gene recapitulates the mutant phenotype.

(a) Schematic of the alb locus showing the position of the T3H48-HHR integration, the target CRISPR site, and the genomic sequences before and after insertion. (b) Relative alb mRNA levels in 36 hpf wild-type and alb^HHR/HHR embryos; n=3 biologically independent samples; Ct values are listed in S1 Table. (c-f) Brightfield images of 36 hpf wild-type (c), alb^b4/b4 (d), alb^HHR/b4 (e), and alb^HHR/HHR (f) embryos. (g) Schematic of the integrated T3H48-HHR in the alb gene, the new CRISPR site created by the insertion, the A>G editing window of the ABE-Umax, and the A>G inactivating mutation in the ribozyme. (h-k) Brightfield imaging (h-i) and sanger sequencing (j-k) of part of the ribozyme and flanking element from 36 hpf alb^HHR/b4 non-injected (h,j) or injected with ABE-Umax mRNA and a sgRNA that inserts an inactivating mutation in the T3H48-HHR (i,k). The proportion of embryos matching the image shown is indicated in the top right corner of each image. The diagram indicates the Anterior-Posterior (A-P) and Dorsal-Ventral (D-V) axes.