Figure 1.

Generation of knock-out alleles of αB-crystallin genes by CRISPR/Cas9. A, alignment of the DNA sequences of the mutant alleles of αBa and αBb. B, electrophoresis of the genomic DNA of the adult zebrafish amplified by PCR: αBa^35bpDEL on 3% agarose gel and αBb^10bpDEL on 4% agarose gel. The genotyping of αBb^10bpDEL allele was confirmed by digestion with restriction enzyme BseLI. C, Western immunoblot of the water-soluble and water in-soluble protein fraction of the excised lenses of the adult zebrafish. After transfer on nitrocellulose membrane, proteins were probed with polyclonal antibody against zebrafish Cryaa (αA) or zebrafish Cryaba (αBa). D, αB-crystallin iTRAQ ratios in the lens of both αBa^−/− and αBb^−/− adult fishes. The left panel shows iTRAQ ratios of αBa and αBb in mutant animals compared with wild-type animals in the lens WSF, and the right panel shows the same from the WIF.