AFLP Markers Linked To Turnip Mosaic Virus-Resistance Gene In Cabbage (Brassica Oleracea Var.capitata)

Cabbage (Brassica oleracea var. capitata ) is a worldwide vegetable crop belonging to the Brassica of family Cruciferae, and it originates from the Mediterranean Coast. The incidence of virus disease has caused tremendous effect and economic loss on the production of cabbage. TuMV (Turnip Mosaic Virus) is one of the major pathogens of virus disease in cabbage, which is introduced into plant cells in the non-persistent transmission mode during aphid probing or feeding. The application of the pesticides to prevent the spread of TuMV is less effective. So breeding the disease-resistant varieties is the economic and effective method to control virus spread. During the transfer of disease-resistance gene by traditional breeding, it needs inoculation to identify the resistance, which is time consuming and labor intensity. And there are some problems such as the loss of objective traits and the low efficiency during the gene transferring.The development of MAS (Marker-Assisted Selection) provides a rapid and effective method for the plant disease-resistance breeding. AFLP (Amplified Fragment-Length Polymorphism) has application extensively in the tagging of target gene for its advantages of high polymorphisms and reproducibility.In this study, the female parent 01-16-5-7, which is a TuMV-susceptible inbred line in cabbage, and the male parent 20-2-5 which is a TuMV-resistant inbred line were crossed to generate the F2 segregation population. The parents and each individual in F2 population were inoculated with the TUMV-C4, a major pathogen in cabbage and then scored for either TuMV-resistant or TuMV-susceptible by the ELISA (Enzyme-Linked Immunosorbent Assay). According to the results of disease-resistance assay, the most TuMV-resistant and TuMV-susceptible individuals in F2 population were chosen to construct two pairs of pools-resistant and susceptible respectively by the method of BSA (bulked segregation analysis), in order to find the molecular marker linked with the TuMV-resistant gene with AFLP techniques in cabbage. The main results were as follows:1. In the closed greenhouse, the seedlings were inoculated with TUMV-C4 artificially and assayed. According to the results of ELISA, the observed ratio of resistant individuals to susceptible individuals was 3.10:1, which was in accordance with 3:1 of Medelian segregation law by the Chi-square (x2) test. It showed that the TuMV-resistance was dominant and monogenic in this population.2. By the method of BSA, two pairs of pools, resistant pool and susceptible pool, were constructed with the pre-amplification product of the most resistant individuals and the most susceptible individuals in F2 population, and total 128 primer combinationswhich consisted of 8 EcoRI primers and MseI primers were used to screen the AFLP markers linked with the TuMV-resistant gene in cabbage by the method of silver staining. As a result, the differences were found between the resistant pools and the susceptible ones, which amplified with the primer combination E24M61. And the marker was named as E24M61-530.3. The amplifications of DNA from parents and individual plants in F2 population with the primer combination E24M61 were repeated three times steadily. In the 172 individuals, there were only 23 crossover plants, which showed that the AFLP marker was linkage with the TuMV-resistant gene in cabbage. The genetic distance between AFLP marker and resistance gene was 14.44cM calculated by the maximum likelihood estimation, and through the adjustment of Kosambi map function.