| Cotton Verticillium wilt, caused by Verticillium dahliae, is a typical vascular systemic disease, and is also the most common, difficult to prevention and treatment disease in China, and this disease also restricts the yield and quality of cotton directly. In this study, the vector containing GFP gene was transformed into Verticillium dahliae, and the strain was used to infect cotton materials dealed with reciprocal graft technique, the invasion and dynamic expansion process of pathogen in cottons were observed by fluorescence microscope and confocal laser microscope, and the distribution of pathogen were investigated by real-time PCR in order to analyze the differences of pathogen colonization and expansion process in sensitive and resistance cottons, so as to further clarify the mechanism of cotton against to Verticillium wilt.The CaCl2-PEG mediated transformation method was used to transform the vector cotaining GFP gene into Verticillium dahliae VD-25, and obtained81transformant strain. According to biology character, fluorescent signal, genetic stability and pathogenicity, we obtained transformant strain G25-35. The biology character and pathogenicity of this transformant strain was similar to the original strain, and this strain had not only strong fluorescent signal but also good genetic stability. So this named G25-35strain was used to infect cotton.The GFP-labeled Verticillium dahliae was used as a reporting system to infect resistant Hai7124and susceptible Jill cotton materials, the materials taken at different time points were observed under the laser confocal microscope and fluorescence microscope, the process of Verticillium dahliae invasion and dynamic expansion in sensitive and resistance cottons were recorded. The results show that the pathogen can infect either resistant or susceptible cotton roots with no difference, and there was no significant difference of the initial infection sites among different cotton types, but the conidia on the sensitive cotton roots germinat earlier than the conidia on the resistance cotton roots, and the pathogen infection rate in sensitive cotton vascular tissues was faster than in resistance cotton. Furthermore, cotton lines with different resistance to verticillium wilt were selected, and reciprocal graft were made between resistant and susceptible cottons that own1-2leaves to obtain the grafting cottons:reciprocal grafting types resistant/susceptible and susceptible/resistant, self-grafting types susceptible/susceptible and resistant/resistant, the ungrafted cottons as control check. The grafting combinations were inoculated, the data of cotton Verticillium wilt were investigated, real-time quantitative PCR was developed to quantify Verticillium dahliae population in different parts of cottons. The results indicated that the resistance was not significantly different between self-root cotton and their self-grafted cotton. The resistance of resistant/susceptible and susceptible/resistant grafting combinations was stronger than those of susceptible/susceptible combination, and weaker than resistant/resistant combination. Pathogen population are variant in different cotton varieties and different parts of cottons. There was significantly higher infection coefficient in different parts of susceptible Jill than in the other cultivars, and the IC value increase upward. However, the fungi quantification was the highest in the hypocotyls of resistant Hai7124and Pima90, and decrease upward. Resistant cotton genotypes either as stock or scion can inhibit the Veiticilliun dahlia, which indicated that the whole organisms of resistant cotton showed resistance to Verticillium wilt, and the resistance ability of underpart stem was stronger than upper stem. |
