Identification of Meloidogyne incognita, M-java']javanica and M-Arenaria using sequence characterised amplified region (SCAR) based PCR assays

Three randomly amplified polymorphic DNA (RAPD) markers, OPA-12(420). OPB-06(1200) and OPA-01(700), species specific to the root-knot nematode species Meloidogyne arenaria, M. incognita and M. javanica respectively, were identified. After sequencing these RAPD-PCR products. longer primers of 18 to 23 nucleotides were designed to complement the terminal DNA sequences of the DNA fragments. This resulted in three pairs of species specific primers that were used to amplify the sequence characterised amplified regions (SCARs). The developed sets of SCAR primers were successfully used in straightforward, fast and reliable PCR assays to identify M. incognita. M. javanica and M, arenaria. The length variant SCAR markers can be amplified from DNA from egg masses, second stage juveniles and females. This species identification technique is therefore independent of the nematode's life cycle stage. Moreover the SCAR-PCR assay was successfully applied using DNA extracts from infested plant material. The method has potential to be optimised for routine practical diagnostic tests facilitating the control of these economically important pest organisms.