Development of a low-density SNP chip for pedigree identification in Portunus trituberculatus

Portunus trituberculatus is an important marine aquaculture crab species in China, widely distributed in the Bohai Sea, Yellow Sea, East China Sea, and South China Sea. Although the aquaculture production and area have been increasing year by year, the coverage of improved breeds remains low, and there is an urgent need to develop new varieties that are adapted to different farming regions. In aquaculture breeding, family information is crucial for accurately estimating genetic parameters, breeding values, and genomic selection. However, due to the large number of individuals and limited marker techniques, family management and information collection face significant challenges. SNP chips, as a high-throughput and low-cost genotyping platform, have been successfully applied in family identification in other species; however, no such application has been reported in P. trituberculatus. This study aimed to design a low-density SNP chip for P. trituberculatus and establish a high-accuracy, cost-effective method for pedigree identification. Based on 298 whole-genome resequencing datasets, high-quality SNP loci were screened from the reference genome with filtering criteria on sequencing depth, minor allele frequency (MAF), and missing rate. Their potential utility for parentage analysis was further evaluated through population genetic approaches. A cGPS chip was then designed using the selected SNP loci, and it was applied for family samples genotyping and kinship analysis combined with the KING-robust algorithm. In total, 977 high-quality SNPs distributed across 53 chromosomes were selected, with an average inter-marker distance of 935 512.44 bp. The mean sequencing of these loci was 13.57×, and the average MAF was 0.21, as well as the average missing rate was 0.11. Population structure analysis demonstrated that these selected SNPs clearly distinguished wild populations, cultured populations, and full-sib families. The designed low-density cGPS chip achieved a locus call rate of 99.48% and a concordance rate of 99.55% across duplicate samples. Genotyping of 45 individuals from eight families revealed an offspring detection rate of 91% and a family assignment accuracy of 100%. Overall, this study successfully developed low-density SNP chip with accurate and stable genotyping performance. It could meets the requirements for pedigree identification and parentage tracing in breeding programs. This chip, along with the associated pedigree analysis framework, provide an important molecular tool and technical foundation for the selective breeding of P. trituberculatus, thereby facilitating the development of enhanced genetic strains.