CRISPR/Cas9 Disruption of oca2 and tyr Genes Alters Pigmentation in Zebrafish

Researchers have used the CRISPR/Cas9 gene-editing system to disrupt two pigmentation genes, tyrosinase (tyr) and oculocutaneous albinism II (oca2), in zebrafish (Danio rerio), producing visible changes in the coloration of edited fish. The study, published in BMC Biotechnology on 16 October 2026, describes the design, synthesis and validation of gene-editing constructs targeting these two genes, which are known to be associated with pigmentation.

Zebrafish holds particular importance in the ornamental fish industry, where it is the most significant species and is available in numerous color varieties. Targeted disruption of genes has already been achieved successfully in several model and non-model fish species, and CRISPR/Cas9 has recently been used to disrupt pigmentation genes in a number of fish species, including zebrafish itself. However, the authors note that few reports to date have described the use of CRISPR/Cas9 to disrupt the tyr and oca2 genes specifically in Danio rerio, which motivated the present work.

The experimental workflow began with the design and synthesis of guide RNA (gRNA) constructs specific to the tyr gene, designated T1, and the oca2 gene, designated T2. In parallel, Cas9 mRNA was synthesized through in vitro transcription.

Both the Cas9 mRNA and the gRNA constructs were validated before use, with gel electrophoresis and sequencing confirming the successful synthesis and validation of the T1- and T2-targeting Cas9 mRNA and gRNA constructs.

Once validated, the Cas9 mRNA and gRNA mixture was co-microinjected into zebrafish embryos at the one- to two-cell stage. The embryos were obtained through natural spawning and were subsequently reared under controlled conditions.

Phenotypic studies of the F0 and F1 generations of mutants showed that melanophores were reduced when the T1 and T2 genes were disrupted, leading to visible pigmentation alterations. The observation of altered pigmentation across both the F0 generation and the F1 generation indicates that the effect was carried through to the next generation of fish.

The authors are careful to note an important limitation of the current study: further studies are required at the molecular level to confirm the mutation by genotyping. In other words, while the phenotypic evidence of altered pigmentation is consistent with successful disruption of the target genes, the genetic changes themselves have not yet been confirmed by sequencing the edited animals’ DNA, and the paper does not report such genotyping data.

Beyond the immediate results, the researchers suggest a broader application for their work. The validated gene-editing constructs developed in the study may serve as useful tools for targeted pigmentation modification in other ornamental fish species, thereby supporting the development of high-value ornamental strains and advancing genetic improvement programs in aquaculture.

For the ornamental fish sector, the ability to deliberately modify pigmentation genes could support the development of strains with novel or enhanced coloration, provided that molecular confirmation of the edits by genotyping is established in future work. The present study documents the construction and validation of the editing tools and reports the resulting phenotypic changes in F0 and F1 zebrafish, with genotyping confirmation identified as the necessary next step.

Subject of Research: CRISPR/Cas9 disruption of the tyr and oca2 pigmentation genes in zebrafish

Article Title: CRISPR/Cas9 induced disruption of oca2 and tyr genes associated with pigmentation in Danio rerio

Article References: Kumari, K., Iqbal, G., Priyanka, S. S., Nidarshan, N. C., Rasal, K. D., Sonwane, A. A., Brahmane, M. P., Bedekar, M. K., Sawant, P. B., & Goswami, M. (2026). CRISPR/Cas9 induced disruption of oca2 and tyr genes associated with pigmentation in Danio rerio. BMC Biotechnology. https://doi.org/10.1186/s12896-026-01239-6

Image Credits: AI Generated

DOI: 10.1186/s12896-026-01239-6

Keywords: CRISPR/Cas9, zebrafish, Danio rerio, tyr gene, oca2 gene, pigmentation, melanophores, gene editing, ornamental fish, aquaculture, genetic improvement, BMC Biotechnology