The Research On The Disease Resistance Mechanism Of The CDPK And Rboh Signaling Pathways And The Selection Of Disease Resistant Varieties

Calcium ion is the second messenger which is commonly found in plants.It plays an important role in plant growth and development,regulation of secondary metabolism and adaptation to the environment.CDPK is a kind of calcium sensing protein with the calcium ion effector structure and the protein kinase effector structure domain.So it is not only the calcium ion effector but also has the enzyme activity.There are many studies about the function of CDPK,such as the growth and the development of plants,in response to biotic and abiotic stresses.Turnip which is a biennial herb of the cruciferous family is the traditional crop of the Qinghai-tibet.The high radiation and low temperature of the Qinghai-tibet result in the low temperature,ultraviolet and disease and insect pests resistance.However,plant diseases and pests are serious.Therefore,it is very important to study the disease resistance mechanism.In this paper,the following three studies were carried out by means of physiological biochemistry and transcriptome,by combining the different resistance varieties.The phylogenetic analysis of the CDPK gene family;Screening of the different resistant varieties;The role of CDPKs in the resistance of powdery mildew.The results are as follows: 1)The phylogenetic analysis of the CDPK gene familyCalcium-dependent protein kinases(CDPKs)as crucial sensors of calcium concentration changes play important roles in responding to abiotic and biotic stresses.In this study,55 BrrCDPK genes,which were phylogenetically clustered into four subfamilies,were identified.Chromosome locations indicated that the CDPK family in turnip expanded by segmental duplication and genome rearrangement.Moreover,gene expression profiles showed that different BrrCDPKs were expressed in specific tissues or stages.Transcript levels of BrrCDPKs indicated that they were involved in abiotic and biotic stresses and that paralogs exhibited functional divergence.Additionally,we identified 15 Rboh genes in turnip;the results of yeast two-hybrid analysis suggested that BrrRbohD1 interacted only with BrrCDPK10 and that BrrRbohD2 interacted with BrrCDPK4/7/9/10/17/22/23.Then we hypothesized that CDPK genes could regulate the activity of RbohD1/2 inducing the accumulation of ROS and the closure of stomata to prevent further infection of pst DC3000.2)Screening of different resistant varieties of turnipWe found that there was difference in the resistance of different cultivars to powdery mildew by homogenous garden experiment.B03 and B14 were regarded as infected and resistant varieties because B03 susceptible to powdery mildew and B14 not easily infected with powdery mildew.The pathogen was identified by morphological and molecular biological methods.Then the physiological and biochemical tests were carried out on three different samples of the two cultivars.The results show that B03 can accumulate more reactive oxygen species,while B14 has stronger ability to remove ROS.We speculate that the excess ROS produced by B03 may cause secondary damage to plants.The results of phytohormone which were closely related to plant defense showed that there was no significant change about the concentration of SA,JA and ET in B03,while the three hormones in B14 showed significant up-trend.Therefore,the pathogen defense process of plant hormone may be one of the factors of B14 resistance.3)The expression analysis of CDPKs in the resistance of powdery mildewIn order to furture study the molecular mechanism of disease resistance,transcriptome analysis was used to explore the expression differences of the CDPKs between the two species at different inoculation time.Transcriptome data suggest turnip CDPKs and RbohD expressed differently in B03 and B14.Among them the starting values of 28 CDPK gene in B14 were higher than that of B03,and 28 common raised genes,8 and 15 specific raised genes in B03 and B14 respectively.RbohD did not increased significantly in B03,but increased in B14.Further analysis revealed that there were no genes interacted with RbohD in the specific up-regulated genes of B03,whereas there were 5 genes in B14.Therefore,we suggested that CDPKs were activated to regulate RbohD accumulating ROS to defense pathogens.And B14 can maintain the balance of ROS by antioxidant system to effectively control powdery mildew infection.