MSc

The investigation on “Molecular markers for bacterial wilt resistance in mapping populations of tomato” was undertaken at the Centre for Plant Biotechnology and Molecular Biology and Department of Olericulture, College of Horticulture, Vellanikkara during the period 2006-2008 to detect trait related markers linked to bacterial wilt resistance. Anagha was used as the resistant parent. DVRT-1 and Pusa Ruby were used as the susceptible parents.F1 plants of both the crosses were found to be susceptible when grown in wilt sick field. F1 plants were raised in sterile media in pots and were selfed to produce F2 population of both the crosses.
F2 populations were raised in a bacterial wilt sick plot. F2 population of both the crosses, after transplanting into bacterial wilt sick plot, showed wilt symptoms. The symptom started as leaf drooping followed by complete wilting and death of the plant. Bacterial ooze test was performed to confirm the infection by Ralstonia solanacearum. Among the two crosses evaluated, wilt incidence was more in F2 population of the cross Anagha x DVRT-1. In the F2 population of the cross Anagha x DVRT-1,out of the 200 plants ,28 plants were observed to be resistant to bacterial wilt and 157 plants were found to be susceptible ,confirmed with ooze test and 15 plants wilted for reasons other than bacterial wilt.

In the F2 population of the cross Anagha x Pusa Ruby, out of the 200 seedlings transplanted, 62 were observed to be resistant to bacterial wilt and 130 plants were found to be susceptible. Plants wilted for reasons other than bacterial wilt were eight in number. Genomic DNA of parents, F1 and F2 populations was isolated, purified and subjected to RAPD assay for molecular characterization. The protocols suggested by Rogers and Bendich (1994) and kit from Chromous Biotech Pvt Ltd. were used for DNA isolation.

The five genotypes i.e. resistant parents, susceptible parent, F1, F2 susceptible bulk and F2 resistant bulk, differing in reaction to bacterial wilt disease were characterized using 18 selected primers from OPF, OPAZ, OPS, OPAH, and OPY series. Though amplification was obtained with all the 18 primers, polymorphism was produced by only one primer OPS 16, in only one cross i.e. Anagha X DVRT-1.RAPD assay could not identify the specific marker for resistance/susceptibility to bacterial wilt disease except with OPS 16 primer, which gave a unique band of size 1.3 kb in Anagha, Anagha X DVRT-1 F1, Anagha XDVRT-1 F2 susceptible bulk and Anagha X DVRT-1 F2 resistant bulk, but was absent in DVRT-1.Polymorphism was not observed between Anagha and Pusa Ruby in the RAPD assay done with the 18 selected primers. So further study of segregation of F2 population was carried out only in F2 population of Anagha X DVRT -1.
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Whole F2 population of the cross Anagha x DVRT-1 was analyzed individually with the primer OPS 16. DNA of all the resistant plants showed 1.36 kb band. In the case of susceptible plants, a segregation of polymorphic band was observed. All this suggests that the gene for resistance to bacterial wilt in Anagha is recessive in nature and the polymorphic band produced by the primer OPS 16 could be related to bacterial wilt resistance.

The recessive nature of the resistant gene is also evident from the fact that all the F1 plants succumbed to bacterial wilt, when they were grown in bacterial wilt sick plot. All these can be confirmed only by analyzing backcross population or F3 population of Anagha x DVRT-1.So future line of work is to develop backcross population or F3 population, and detect whether the polymorphic band given by the primer OPS 16 is related to bacterial wilt resistance or not.