Preliminary Construction Of F₁ Mapping Population And The Frame Molecular Linkage Map Of Mei Flower

Mei flower was native to China, and plays an important role in gardening of China. At present, the studies on mei flower included germplasm resource, breeding, molecular biology and so on. Yet, there were no published articles about linkage map of mei flower. The research based on molecular biology needed further development.In this research, the frame molecular linkage map of mei flower was constructed according to the method of "double pseudo-testcross", with the RAPD and SSR data obtained from the F₁ population of 'Xuemei'×'Fenpi Gongfen'. The construction of the map aimed at settling a foundation for application of molecular markers, cloning the genes that controlling important traits of mei flower and promoting the development of genomics of mei flower. The main results were as follows:1. Construction of F₁ populationA F₁ population of 43 individuals from a cross between 'Xuemei' and 'Fenpi Gongfen' was constructed. The effects of low temperature treatment and wiping off the endocarp on breaking the dormancy of the seeds were studied.2. Optimization of SSR reaction systemThe factors which affected the SSR amplification of mei flower were studied, using 'Xuemei' and 'Fenpi Gongfen' as the template DNA. In the 20μl reaction system of SSR, the optimal concentration of primers, template DNA, dNTP, Mg²⁺, Taq DNA polymerase was 0.2μmol/L, 60ng, 150μmol/L, 2.0mmol/L, 1U, respectively. PCR reactions were performed by an initial denaturation for 1 min at 94℃followed by 35 cycles of 45s at 94℃, 45s at 55℃to 58℃, 2min at 72℃; and a final extension of 4min at 72℃.3. Parenthood of F₁ populationThe parenthood of 43 individuals of F₁ population was identified by RAPD and SSR. 39 and 25 individuals were confirmed to be true hybrid by the methods of RAPD and SSR, respectively. Because of the small number of SSR markers, 39 individuals identified by RAPD were selected to construct the linkage maps.4. SSR segregation analysis5 in 32 pairs of SSR primers were selected for different amplification between parents. 20 polymorphic loci were obtained. Each polymorphic loci was tested byχ² analysis for goodness of fit (P＜0.05) to the Mendelian segregation ratios. For 'Xuemei', three loci were fit to the 1:1 segregation ratio, and for 'Fenpi Gongfen', there were six. Only one locus was fit to the 3:1 segregation ratio. 5. RAPD segregation analysis52 in 440 RAPD primers were selected for their significant difference between parents. 78 steady and repeatable polymorphic loci were obtained, with an average of 1.5 for each primer. For 'Xuemei', 23 loci were fit to the 1:1 segregation ratio byχ² analysis, and for 'Fenpi Gongfen', there were 34 loci. Six loci were fit to the 3:1 segregation ratio.6. Construction of frame molecular linkage maps of 'Xuemei' and 'Fenpi Gongfen'The maps of 'Xuemei' and 'Fenpi Gongfen' were constructed by using RAPD and SSR markers, respectively. The map of 'Xuemei' with the length of 366.4cM included 17 markers arranged to 5 linkage groups, and the average distance was 22.9cM. The map of 'Fenpi Gongfen' with the length of 556.6cM included 26 markers arranged to 5 linkage groups, and the average distance was 22.3cM.