Genome-Wide Development and Validation of KASP-Based SNP Markers in Neophocaena asiaeorientalis asiaeorientalis

The Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis) is the only extant freshwater cetacean species inhabiting the Yangtze River in China. Facing significant threats from habitat degradation and human activities, this critically endangered species requires urgent conservation efforts. A thorough understanding of its genetic diversity is fundamental for informing effective conservation strategies. To address the need for stable and reliable molecular markers, this study aimed to develop and validate a set of highly polymorphic single nucleotide polymorphism (SNP) markers. Candidate SNPs were initially identified based on chromosome-level genome and whole-genome resequencing data, yielding a total of 1070 candidate loci. Following the principle of even distribution across chromosomes, 50 SNPs were randomly selected for Sanger sequencing validation, from which 35 polymorphic SNPs were preliminarily confirmed. These 35 SNPs were subsequently genotyped using the Kompetitive allele-specific PCR (KASP) assay, which successfully validated 19 highly polymorphic markers. Genetic diversity analysis using these SNPs successfully captured moderate-to-high polymorphism in the Poyang Lake population, demonstrating the capability of these markers in detecting genetic variation. The SNP markers developed in this study show promising applicability for genetic diversity assessment in the Yangtze finless porpoise. Further validation across broader geographic and demographic samples will be essential to confirm their potential efficiency for detecting spatial heterogeneity and informing region-wide management strategies.