Identification of sex in spiny gourd (Momordica dioica) using RAPD marker analysis and its validation in four F1 progenies

Momordica dioica is an important summer vegetable crop in India and is dioecious. DNA bulks of six male and six female accessions of M. dioica were subjected to random amplified polymorphic DNA (RAPD) marker analysis using 40 primers, of which 22 primers responded for amplification. Of these, ten primers generated reproduction type-specific amplification of DNA fragments at the bulk level. This number was reduced to three RAPD primers when re-tested, individually, with six male and six female accessions. One random decamer primer, OPA-01, produced two unique amplified fragments in all female plants, while two other primers, OPB-04 and OPC-08, generated distinct amplified fragments in both male and female plants. Three amplified fragments (OPB-04(430), OPC-08(1135), and OPC-08(578)) were unique to all six male accessions, while five amplified fragments (OPA-01(1560), OPA-01(880), OPB-04(880), OPB-04(800), and OPC-08(1230)) appeared uniquely and uniformly in all six female accessions. Southern hybridisation analysis was performed using five of these sex-specific amplified fragments (OPA-01(880), OPB-04(880), OPB-04(430), OPC-08(578), and OPC-08(1230)) as probes with Eco RI and Hind III double-digested DNA of all male and female accessions. Southern hybridisation analysis with the OPC-08(578) probe showed a unique hybridisation pattern for all six male accessions, as with the respective RAPID marker, whereas the other probes showed complex hybridisation patterns. Further, inheritance studies also confirmed the presence of a homologous restriction fragment targeted by this probe in F-1 progenies, as expected. Hence, OPC-08(578) could be used as a marker to ascertain the sex of M. dioica plants at an early stage of development.