Development of snp-dependent KASP system for the identification of economically important root-knot and cyst nematodes in potato

Potato is one of the basic foods for feeding the world's population. In addition to its fresh consumption, potato is also used as animal feed, seed and industrial starch. Cultivation of potato which have economic importance are many pest and diseases. Globodera rostochiensis, Globodera pallida species from potato cyst nematodes; Meloidogyne hapla and Meloidogyne chitwoodi species from root-knot nematodes are the pests of potato plants. These species cause loss of productivity in potato production, decrease in market value and limitation of production areas worldwide. These species need to be identified accurately and quickly in order to take quarantine measures and determine management strategies. In this study, KASP (Kompetitive Allele-Specific PCR) technique based on single nucleotide polymorphisms (SNP) was used to identify the species G. rostochiensis, G. pallida, M. hapla and M. chitwoodi. SNPs were determined for each species using the available DNA sequences of these species from GenBank. KASP-Gr, KASP-Gp, KASP-NMhp and KASP-Mch for G. rostochiensis, G. pallida, M. hapla and M. chitwoodi were developed, respectively. These primers were able to successfully distinguish G. rostochiensis, G. pallida, M. hapla, M. chitwoodi species from each other and from other plant parasitic nematodes. These species were identified using DNA obtained from different biological stages. Globodera rostochiensis and G. pallida species were identified from one cyst using KASP-Gr and KASP-Gp primers. Studies conducted at the larval level showed that >8 juvenils DNAs for the G. rostochiensis species and 1 juvenile DNA for the G. pallida species were sufficient to identify these species. KASP-NMhp primers developed to identify M. hapla species were found to work successfully in 1 juvenile DNA. It was determined that the KASP-Mch primers developed to identify M. chitwoodi species worked in >20 juvenils DNAs. In addition, KASP-Mch primers successfully identified DNAs obtained by direct isolation from infected potato tuber tissue with M. chitwoodi. The developed primers can be used accurate and quick identification of these species in many laboratories, especially quarantine laboratories.