New Materials With High Resistance To Clubroot Were Created By Interspecific Hybridization Polymerization Of Different Resistance Sites In Brassica

Clubroot disease is a worldwide soil-borne disease,which is extremely harmful to cruciferous crops.Once infected by plasmodiophora brassicae Woron,the yield will be reduced if the disease are light,and will not be harvested if the disease are heavy.At present,the incidence area of clubroot in China is increasing year by year,and it has become the main disease of Brassica crops such as Chinese cabbage（Brassica rapa）,cabbage（Brassica oleracea）and Brassica napus,which had caused great losses to their yield and quality.At present,prevention and control measures to clubroot include mainly agricultural control,biological control and chemical control and so on,but effect are poor,and breeding resistant varieties is considered to be an effective control means.In China,most disease-resistant varieties of Brassica vegetables in production have fewer resistance sites,mainly resistant to Clubroot 4.However,plasmodiophora brassicae has many physiological races,which greatly increases the possibility of losing the resistance of disease-resistant varieties in a short period.Therefore,the creation of Brassica anti-clubroot materials with long-lasting resistance will be of great significance to the clubroot-resistant breeding of Brassica vegetables.The purpose of this study was to create new Brassica vegetable materials with high resistance by interspecific hybridization combining embryo rescue technology and molecular marker-assisted selection（MAS）.The main results are as follows:1.Cabbage 9LQ（CRb）,chinese cabbage 8574（CRd）and turnip ECD4（Pb Ba8.1）were hybridized combining embryo rescue technology.The results showed that 9LQ×ECD4 obtained 6 viable young embryos.Natural hybridization between Chinese cabbage 8574 and turnip ECD4 can produce seeds normally.OA and YA primers are used to detect CRb,and A08-300 primer are used to detect Pb Ba8.1,and Yau389primer are used to detect CRd;Two materials with resistance sites of CRb and Pb Ba8.1 were screened out from 6 9LQ×ECD4 materials.Out of 9 8574×ECD4materials,four materials with resistance sites of CRd and Pb Ba8.1were screened out from 9 8574×ECD4 materials.The morphology and pollen viability of the two resistant loci materials were analyzed.The hybrid offspring were different from their parents.The stamens of 9LQ×ECD4 were shriveled and could not produce pollen,and the fertility of 8574×ECD4 and their parents was normal.Resistant identification of 9LQ×ECD4 distant hybrid progeny showed it is resistant to race 4 in Wulong District of Chongqing and race 11 in Chengdu,Sichuan Province.2.On the basis of obtaining two resistance sites materials,six cross combinations were hybridized with Z3(CRb^kato)to create three-resistance site materials,and 18surviving young embryos were obtained by distant hybridization combining with embryo rescue technology,namely,3R-1～3R-9 of Z3×（8574×ECD4）combinations,and 3R-10～3R-15 of（8574×ECD4）×Z3 combinations,and 3R-16～3R-18 of（9LQ×ECD4）×Z3 combinations.After natural fruiting in the field,nine surviving seedlings were obtained from the two combinations,which were 3R-19～3R-22 of Z3×（8574×ECD4）combinations and 3R-23～3R-27 of（8574×ECD4）×Z3combinations.3.Through molecular marker-assisted selection,two materials with three resistance sites were screened out from 18 surviving young embryos;Two materials with three resistance sites were screened out from 9 natural seedlings,and four materials with three resistance sites were obtained,which were 3R-9,3R-10,3R-19and 3R-26 respectively.Morphological analysis showed that the three resistance sites materials were true hybrids,and pollen viability analysis showed that the pollen fertility of 3R-9,3R-10 and 3R-19 were all 100%,and the pollen fertility of 3R-26was partially sterile,with a fertility of 26.15%.Resistance of 3R-9 and 3R-10 which were inoculated with Chongqing Wulong No.4 and Sichuan Chengdu No.11 showed immunity.This,indicated that the created three-resistance site materials resitant to No.4 and No.11 physiological races.4.Three resistance sites materials 3R-9,3R-10,3R-19 and 3R-26 were hybridized with single resistance site material 9LQ（CRb）to create four resistance sites materials.Among the eight combinations,3R-9×9LQ combination obtained five viable young embryos,and 3R-10×9LQ combination obtained one viable young embryo,3r-26×9LQ combination obtained two viable young embryos,and obtained eight distant hybrid progeny,4R-1～4R-8,.When 9LQ was used as female parent,viable young embryos all combination were not obtained,which indicated that reciprocal crossing had a great influence on the compatibility of distant hybridization.Four pairs of primers B0902,Yau389,A08-300 and OA were used for molecular marker-assisted selection of 4R-1～4R-8,and one material with four resistance sites（4R-6）,and two materials with three resistance sites materials,four materials with two resistance sites,one material with one resistance site were successfully screened.The morphological analysis of 4R-6 shows that its leaves are wavy and grow healthily,and the leaves begin to have the ball formation.5.Four resistance site materials（4R-6）was inoculated with Plasmodiophora Brassicae from Wulong in Chongqing city,Fuling in Chongqing city,Wanzhou in Chongqing city,Chengdu in Sichuan province,Guangyuan in Sichuan province,Panzhihua in Sichuan province,Yichang in Hubei provinceand Enshi in Hubei province,and 4R-6 showed immune to Plasmodiophora Brassicae from Wulong in Chongqing city,Fuling in Chongqing city,Wanzhou in Chongqing city,Chengdu in Sichuan province,Guangyuan in Sichuan province and Enshi in Hubei province,and showed tolerant to Plasmodiophora Brassicae from Panzhihua in Sichuan province,and show susceptible to Plasmodiophora Brassicae from Yichang in Hubei province,moreover,its disease index（37.5）was lower than control（60.71）.The results showed that 4R-6 with four resistance sites material showed great resistance to Clubroot in many areas.