Exploration of enhancing hypoxia tolerance in Nile tilapia（Oreochromis niloticus） through setd7 gene editing

This study aims to explore the function of the methyltransferase gene setd7 in hypoxia tolerance in fish, using Nile tilapia (Oreochromis niloticus) as the experimental model. By employing gene editing technology, we constructed setd7 gene mutation lines to assess their physiological responses and tolerance capabilities in low oxygen environments, providing theoretical basis and experimental materials for molecular breeding of hypoxia tolerant fish and research on the functional mechanisms of methyltransferases. Using CRISPR/Cas9 gene editing technology, specific knockout targets were designed for the conserved amino acid region of the second exon of the setd7 gene in O. niloticus, and microinjection was performed on one cell fertilized eggs. The gene editing efficiency was verified through T7E1 enzyme digestion experiments and sequencing analysis, successfully obtaining the setd7^+/− mutation line with a -23 bp base deletion. Further, one month old F₁ generation tilapia were subjected to hypoxic stress treatment to observe their hypoxia characteristics and survival time, assessing their hypoxia tolerance. The experimental results showed that the setd7^+/− mutation line exhibited a loss of function of the SETD7 protein due to a frameshift mutation, with a significantly reduced expression level of the setd7 gene compared to the wild type. In the hypoxic stress experiment, the setd7^+/− mutant tilapia demonstrated stronger hypoxia tolerance, with the onset of surface swimming occurring significantly later than in the wild type, and a markedly extended survival time in low oxygen environments. This indicates that the mutation of the setd7 gene significantly enhances the hypoxia adaptation ability of tilapia. This study found that mutations in the methyltransferase gene setd7 can significantly improve the hypoxia tolerance of Nile tilapia, providing important experimental materials and theoretical foundations for molecular breeding of hypoxia tolerant tilapia. Additionally, this research lays a scientific foundation for further exploration of the molecular mechanisms and functional studies of methyltransferases in hypoxia adaptation.