In this study, we demonstrated that lrp6a, a co-receptor in the Wnt signaling pathway, is essential for proper median fin formation and somitogenesis in goldfish. We analyzed the gene’s sequence features and expression patterns in both wen-type and egg-type goldfish, uncovering distinct tissue-specific expression differences between the two varieties. To explore the functional role of lrp6a, we performed CRISPR/Cas9-mediated gene knockout using eight designed single-guide RNAs (sgRNAs), of which four showed effective targeting. Three high-efficiency sgRNAs were selected and co-injected into embryos to achieve complete gene disruption. Morphological assessments and X-ray microtomography (μCT) imaging of the resulting mutants revealed various abnormalities, including defects in the dorsal, caudal, and anal fins, as well as skeletal deformities near the caudal peduncle. These results confirm that lrp6a plays a key role in median fin development and axial patterning, offering new insights into the genetic regulation of fin formation in teleost fish.

The twin-tail phenotype of goldfish represents a striking domestication-associated remodeling of the vertebrate caudal axial system and is classically linked to disruption of Chordin/BMP-mediated dorsal–ventral patterning. Although previous knockdown studies implicated sizzled (szl) in this process, genetic evidence from targeted disruption of endogenous szl loci remains limited. Here, we used CRISPR/Cas9 to mutate conserved coding regions shared by the duplicated goldfish paralogues szlA and szlB in single-tail embryos. Sanger sequencing and ICE analysis showed that szl-sgRNA2 and szl-sgRNA3 efficiently induced indels at both loci, whereas szl-sgRNA1 was ineffective. Across three independent biological replicates, twin-tail-like caudal bifurcation was observed in 44.63–48.19% of szl-sgRNA2-injected larvae, 69.47–79.61% of szl-sgRNA3-injected larvae, and 64.29–76.19% of larvae injected with the sgRNA mixture; szl-sgRNA1-injected larvae remained single-tailed. Calcein staining further revealed separation of distal caudal fin rays and partial splitting of the caudal skeletal complex in szl-edited larvae. qRT-PCR showed selective remodeling of dorsal–ventral patterning genes, including reduced chdA and eve1 expression and increased bmp2 and nog1 expression. These findings provide direct functional evidence that szl regulates median caudal patterning in goldfish and suggest that szl-dependent modulation of the Chordin/BMP network can generate twin-tail-like caudal morphology.

Accurate pedigree reconstruction is essential for genetic evaluation, inbreeding control, and family management in aquaculture breeding programs. In sturgeon, extremely high fecundity and communal rearing during early developmental stages often lead to the loss of family information, making reliable full-sib family assignment a critical challenge. In this study, we developed a machine learning-based framework for full-sib family assignment using simulated genotype datasets and whole-genome resequencing data from Russian sturgeon. Simulation analyses across five machine learning algorithms showed that training set size and marker density were the primary determinants of assignment accuracy. When at least 10 individuals per family were included in the training set, mean identification accuracy exceeded 99% across all evaluated scenarios, and exceeded 99.9% for all methods except XGBoost. In contrast, performance declined when the marker number was reduced to 200. At moderate marker densities (500–1000 markers), performance remained stable, with mean identification accuracy around 99% even when only 3–4 individuals per family were included in the training set. Validation using whole-genome resequencing data (sequencing depth ranging from 9.43× to 11.86×) from 582 individuals representing 19 full-sib families of Russian sturgeon confirmed the simulation findings, with several algorithms achieving assignment accuracies exceeding 99%. These results demonstrate that machine learning provides an accurate and robust approach for full-sib family assignment using genome-wide single nucleotide polymorphism (SNP) data. The proposed framework offers an effective solution for pedigree reconstruction and family identification in sturgeon breeding populations lacking reliable pedigree records.