Development and validation of methylation markers related to low salinity tolerance in Portunus trituberculatus

In order to develop methylation markers related to low salinity tolerance in Portunus trituberculatus, in this study, methylation sensitive high resolution melting curve method (MS-HRM) was applied to screen and validate methylation loci related to low salinity tolerance. A total of 8 methylation markers were developed from the transcriptome database of P. trituberculatus, six of which were significantly related to low salinity tolerance, exhibiting a significant demethylation or methylation pattern in the low salinity condition. They were located in V-type proton ATPase, H⁺/Cl⁻ exchange transporter, succinate dehydrogenase, NAD(P) transhydrogenase, phosphoglycolate phosphatase, NADH dehydrogenase genes, and these markers provided useful tools for marker-assisted breeding of low-salinity-tolerant strain of P. trituberculatus. The low salinity tolerance test and genotyping results of these 6 loci in wild populations and “Ningxiang-1” selective strain showed that the survival rate of “Ningxiang-1” was significantly higher than that of wild populations under low salinity conditions, and the demethylation of loci Pt-M1 and Pt-M2 in the selective breeding population was significantly increased to 100%, which may be beneficial to maintaining the balance of Na⁺ and Cl⁻ under low salinity conditions. This study provides basic data and favorable tools for genetic improvement of low salinity resistance in P. trituberculatus.