| Cucumber powdery mildew is one of the main diseases of cucumber,caused the leaves turn yellow,crisp,dry,serious impact on Photosynthesis of cucumber leaves,decrease the yield of cucumber,cucumber powdery mildew has a short incubation period,infection and epidemic characteristics of frequent and strong annual,and generally in the middle and late growth of cucumber disease spreads quickly.Bring serious economic losses to production.At present,the main means of prevention and control of Cucumber Powdery Mildew in four ways:strengthening field management,chemical control,disinfection greenhouse and breeding resistant varieties.There are both advantages and disadvantages.The traditional positioning resistance breeding to clear the cucumber powdery mildew resistance gene based on the hybridization,backcross breeding,breeding after many generations,the work is difficult.The RPW8 which be found in Arabidopsis,has broad-spectrum resistance to powdery mildew,this gene contains two genes,RPW8.1 and RPW8.2 respectively.It not only has effect on the resistance of Arabidopsis to powdery mildew,but also showed higher resistance to downy mildew and powdery mildew in Arabidopsis,tobacco tobacco mosaic virus and other plant parasitic pathogens.The RPW8.1 protein located around the chloroplasts,it is more potential of resistant than RPW8.2 protein,has now been confirmed that most of its(80%)transgenic lines resistant to downy mildew,powdery mildew resistance small part(30%),about 10%of both transgenic lines resistant to powdery mildew,and resistant to downy mildew.Therefore,we are going to transfer the broad-spectrum disease resistance gene RPW8.1 into Cucumber and study its function in cucumber.On this basis,the use of the cucumber promoter can make the expression of RPW8.1 in cucumber better,first we need cucumber promoter RPW8.1 homeopathic expression in tobacco,further into cucumber.We got these result:1,we use cucumber powdery mildew,Pseudomonas,to analysis of transcription of PAPP2C gene in Cucumber by qRT-PCR.Results showed that the cucumber PAPP2C gene is affected by cucumber powdery mildew and Pseudomonas bacilli.After inoculation with cucumber powdery mildew and Pseudomonas 24 hrs,the transcription level of PAPP2C was significantly increased,which indicated that PAPP2C could be induced by pathogenic bacteria,so we chose the promoter of PAPP2C to start the expression of RPW8.1.2,We selected CsPP2C10PF1 and CsPP2C10PF2 as candidate promoter fragments of RPW8.1,The pCsPP10CY1-eYFP and pCsPP10CY2-eYFP promoter reporter vectors were constructed,and the transient expression in tobacco.We found that the expression of promoter fragment CsPP2C10PF1 and CsPP2C10PF2 were able to start eYFP,but from the start of the CsPP2C10PF1 fluorescence fluorescence see weaker than CsPP2C10PF2 expression,so we chose CsPP2C10PF2 as a promoter of RP8.1.3,We found that CsPP2C 10PF2 could activate the expression of RPW8.1 in tobacco.We connect the RPW8.1 into the pCsPP10CY2-eYFP vector and observe the fluorescence when injected into tobacco.The results show that RPW8.1 can be activated in parallel with the promoter of cucumber PAPP2C.But the specific function we need to verify in the cucumber.So we need to transfer the vector into cucumber.4,We need to construct the genetic transformation system of cucumber,which is the basis for the cucumber PAPP2C promoter to start the RPW8.1 vector into cucumber.In the system we found that the cucumber seeds soaked for 12 h and then stripped of skin faster than the growth of stripped of skin after 30 min soaking;seed disinfection should be the first to use alcohol treatment,after treatment with sodium hypochlorite;treated with sodium hypochlorite,sodium hypochlorite concentration caused by seed contamination in 4%and 8%were not in the induction medium;in 6-BA,mg/L = 3,ABA = 1 mg/L,the highest rate of bud induced Changchun Mici can reach 86.67%;shoot elongation medium for ABA inhibited shoot elongation,so it can not be used,but the concentration of 6-BA should be reduced to 1 mg/L,otherwise it will cause the vitrification of shoots;bacteria infection at the concentration of OD600 = 0.2-0.25,for 20 min,this will not cause bacterial contamination;Research has shown that the higher the sub leaf differentiation,antibiotic resistance is strong,because Changchun Mici re differentiation rate is very high,it is found that the Kan concentration to 200 mg/L can not caused by normal plant chlorosis,so we decided not to use Kan screening positive plants by plant growth after screening.Cucumber downy mildew is one of the main diseases in cucumber production.It has the characteristics of frequent infection and epidemic.Usually,under the right conditions,the disease spreads quickly.Early in the disease,if treatment is not timely,1-2 weeks to spread,will cause the reduction of output,and even failure,crops,limiting the yield of cucumber.Pesticide has been the main method to prevent and cure diseases.From the earliest copper agent to the present,the single agent can not prevent downy mildew of cucumber.So we're looking for a solution to the problem.According to the Ministry of agriculture(SY201102316),a field experiment was conducted to evaluate the efficacy of 70%-in-situ-fat and water dispersible granules in the control of cucumber downy mildew.The foliar spraying method,according to the different effective dose of 70%azoxystrobin fat-Dimethomorph water dispersible granule composite chemicals and drugs and the control of unilateral contrast,eventually found that 70%azoxystrobin fat-Dimethomorph water dispersible granule composite medicament effective content is higher,the lower the disease index and control index is high effective final recommendations 70%azoxystrobin fat-Dimethomorph agent to control cucumber downy mildew dosage is 367.5g/hm2-420g/hm2 times to 3 times is appropriate in the early stage of the disease or according to the forecast before the onset of pesticide. |
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