Abstract: This study used the CRISPR/Cas9 technology to knock out the hbae1.1 gene and obtained the homozygous mutants of hbae1.1 gene. The effect of hbae1.1 gene deletion on hemoglobin production in zebrafish was investigated. The knockout target was designed according to zebrafish hbae1.1 gene, and the corresponding gRNA was prepared, gRNA and Cas9 protein were mixed in a certain proportion and injected into zebrafish fertilized egg at a cellular stage by microinjection. After T7E1 endonuclease digestion, sequencing and sequence comparison, it was detected that the hbae1.1 gene was successfully knocked out. By mating with wild type and internal crossover, the F2 homozygous mutant was successfully obtained. Comparing the o-Dianisidine staining of embryos produced by the homozygous mutant with hbae1.1 gene deletion and wild type, the results showed that the hemoglobin content of the mutant was significantly reduced compared with the wild type. Studies have shown that the deletion of hbae1.1 gene would have a certain effect on the production of zebrafish hemoglobin. At the same time, the study of hbae1.1 gene provided a certain basis for understanding the occurrence and development of fish hemoglobin.

Key words: zebrafish, hemoglobin, hbae1.1 gene, CRISPR/ Cas9, o-dianisidine