Development of a microsatellite marker set for parentage testing in Japanese commercial three-way cross pigs

The objective of the present study was to construct a parentage testing system using microsatellite markers for Japanese commercial pigs. We selected 10 microsatellite markers based on two criteria. One was high probability of paternity exclusion (PE) in Japanese commercial pigs produced by three-way cross with sires of the Duroc breed and Fsub(1) dams between the Landrace and Large White breeds, the other was accuracy and easiness of genotyping. Heterozygosity, PE and the combined PE (CPE) were calculated from allele frequencies of microsatellite markers. The investigated population was consisted of 472 pigs (124 Duroc pigs and 348 Fsub(1) animals between Landrace and Large White). The PE of selected 10 markers (ACR, SW24, SW2429, SVV1550, SW1027, SW1328, SW443, SO316, SWR1921, and SW1263) ranged from 0.512 to 0.688. The CPE were 0.9999 with sires', dams' and offspring's genotypes and 0.9973 with sires' and offspring's genotypes, in commercial three-way crossed (LWD) pigs. Paternity testing in the field was performed on the two sets of samples to evaluate the actual effectiveness. The set 1 consisted of 35 LWD meat pigs and their 18 Duroc sires. The set 2 consisted of 65 LWD meat pigs and 18 Duroc sires without consanguinity. In the set 1, all LWD pigs were unambiguously assigned to one sire. In the set 2, the set of 10 microsatellite markers did not exclude the paternity of 3 pairs between LWD pigs and false sires. The CPE calculated from the set 2 was 0.9986. The 10 microsatellite markers were further refined to be capable of multiplexed PCR, which is suitable for routine parentage testing. We re-designed PCR primers for five microsatellites and improved the PCR condition ; concentration of each primer and PCR cycle. This microsatellite marker set will provide a powerful and efficient tool for parentage testing in Japanese commercial pigs.