Detection Of Molecular Markers Linked To Genes Conferring Resistance To Bacterial Wilt In Tobacco

Tobacco is an important economic crop in our country. Tobacco bacterial wilt (TBW) is a typical vascular disease caused by bacterium Ralstonia solanacearum. The bacterium invades plants from roots and damages the vascular transport tissue through proliferation and a series of biochemical action, causing the plants dehydrated. Once the disease occurs, the whole plant will die. TBW has become a serious disease affecting the yield and quality of tobacco. This study aimed at detecting SSR markers linked to the resistance genes of TBW so as to facilitate the molecular marker-assisted breeding of tobacco varieties resistant to TBW. The main results are as follows:(1) An F₂ population was derived from a cross between a TBW-resistant cultivar Yanyan-97 (as female) and a TBW-susceptible cultivar Honghuadajinyuan (as male) and was planted on pots in greenhouse. Two hundred uniform plants selected from the F2 population were used for inoculation experiment.(2) At the 5-6 leaf stage, leaves with similar size selected and detached from seedlings, and then were inoculated by injection of Ralstonia solanacearum, with injection of steril water as control. Twenty days after inoculation, Yanyan-97 and F1 leaves still remained green and showed resistance without any disease symptum, whereas Honghuadajinyuan leaves exhibited susceptibility with typical disease symptum. Most of F2 inoculated leaves showed resistance. The result suggests that there might be major resistance genes in the population.(3) DNAs were extracted from the two parents using CTAB method. 1152 SSR primers were used to analyze the parents, and 120 (¹0%) polymorphic primers were obtained.(4) Ten most resistant and 10 most susceptible seedlings were selected from the F2 population to construct a resistant DNA pool and a sensitive DNA pool, respectively. According to the principle of bulked segregant analysis (BSA), the polymorphic SSR primers obtained were used to analyze the two pools. Four SSR markers (M45, M69, Tp1244 and Tp1253) were found to reveal polymorphisms between the two pools, same as those between the two parents, suggesting that these markers were probably linked to TBW-resistance genes. M45 and M69 were previously mapped on the same linkage group with very close distance between them. Tp1244 and Tp1253 were newly developed SSR markers with unknown positions. The result suggests that there is at least one resistance gene identified, and will help more precise mapping of resistance genes and finding more tightly linked molecular markers.