| The proportion of edible vegetable oil provided by ourselves is merely40%, which makes the rapeseed become one of the weakest bulk agricultural products in China. There are two predominant factors to restrict the increase of the rapeseed plant scale:the low resistance and low mechanization level. Lodging not only results in a decrease of rapeseed production which ranges from10%-30%and even more than50%, but also affect the quality that leads to a10%to30%decrease of the oil content. It is also an obstacle to mechanization level increasing. Drought is one of the climatic disasters for crop and agriculture production, the proportion of which in the distorted fields had been as high as75%.Thus, it is a very important approach to enhance the resistance of rape to drought and lodging by investigating the regulation and expression mechanism underlying lignin gene in rape stem under drought stress, which could also contribute to rape breeding for new variety that is wide tolerance, high resistance and more suitable for sowing and harvest by machine.We have selected10species (lines) of Brassica napus with stable highest lodging resistance and lowest lodging resistance, to analyze the difference and dynamic changes of those morphology traits of stem, the lignin content and the rate of different lignin monomers in different development stage, by planting them in a shed and treated with drought condition and also set a control with plenty water. We compared the different expression of lignin metabolic pathway genes and revealed the mechanism for lodging resistance in drought condition.The primary results are as follows:1) Relationship between the traits related to lodging resistance and stem breaking-resistance under drought stress treatment. We collected3plants in each species to investigate the plant height, stem diameter, water content, breaking-resistance in four significant development stages of rapeseed (budding stage, early flowering stage, final flowering and podding stage). The dynamic changes for each trait in both treatments were analyzed, and the effects on breaking-resistance were also analyzed by path analysis. The results showed that:(1) there were significant difference in the performance of the four traits under drought stress and the control treatment. In drought stress environment the plants will be shorter, lower water content and smaller stem diameter, but with stronger breaking-resistance;(2) there also have obviously genotype-environment interaction effect those trails during some of the developmental stages;(3) in both environments plant height had largest direct effect on breaking-resistance, and stem diameter also has certain directly determine on breaking-resistance. The remaining path coefficients showed that here maybe other factors influence the stem breaking-resistance besides plant height and diameter.2) The relationships of rapeseed stem breaking-resistance to the ratio of lignin monomers and lignin content in under drought stress. During four stages of the rapeseed development, we took3plants from the drought treatment and control treatment for each species, determined the ratio of lignin monomers and lignin content using the middle portion stem of the plant. The results show that:(1) The rapid accumulation period of the rapeseed stalk lignin was during the final flowering stage to podding stage;(2) Stem lignin content was greatly impacted by environment and different development stages. Stem lignin accumulation was accelerated under the drought conditions. There also obviously genotype-environment interaction effect to the accumulation of stem lignin except budding stage;(3)The correlation analysis of lignin content and breaking-resistance showed that there were highly significant positive correlation between the lignin content and stem breaking-resistance in both environmental conditions. It confirmed that the higher stem lignin content means the stronger mechanical strength, and the breaking-resistance;(4) The H-lignin accounted for10.96%of the total lignin components by average, and there was little changes in different species. The lignin monomers of rapeseed were mainly constituted by S-G lignin, the differences of the lignin monomer S/G ratio were very significant in the various species;(5) During the budding to the podding stage, S/G ratio of the lignin monomer showed an uptrend in two kinds of environments, and it shows highly significant difference between each stages, which showed that the S-lignin increasingly became the main monomer of the total lignin:The average S/G ratio of stem lignin monomer in the drought condition was slightly higher than control group, but the differences of S/G in various developmental stages haven’t reached significant level. (6)In early flowering stage, final flowering stage and podding stage, the S/G ratio of lignin monomer in lodging resistance species was significantly higher than that of lower lodging-resistant species in both treatments. And the S/G ratio of lignin monomer showed a highly significantly positive correlation to the breaking-resistance of stem. It speculated that in rapeseed S-lignin monomer proportion may closely relate to the lodging resistance.3) The expression analysis of key genes in the lignin metabolic pathways under drought stress. In the drought group and the control group, RNA of three plants stem of each species were extracted during four stages of rapeseed development. The expressions of11key genes in lignin metabolic pathways were analyzed by using qRT-PCR methods. The results showed that:(1) in both environments the expression of PALI gene will increase first and then decreased. The peak of expression under drought stress peak delayed under drought stress condition, and drought treatment can make PALI express significantly higher in the final flowering stage than normal environment. Under control treatment, the PALI expression of lodging-resistance species were significantly higher than that of lodging species in the budding stage, the final flowering and podding stage;(2) the expression of PAL2gene increased first and then decreased. There was a peak in the early flowering stage in both environments. In control condition the expression of PAL2was significantly higher than that in the drought stress environment, which was just opposite in the final flowering stage. In the budding stage the expression of PAL2of lodging-resistance species were significantly higher than lodging species, it doesn’t shows significant difference in other stages;(3) the expression of C4H gene increasing from the early flowering stage to podding stage in both environments, and the expression in control condition always higher than the drought condition; the expression of lodging species was significantly higher than lodging-resistant species in budding stage, final flowering stage and podding stage, and in the podding stage the difference expression became maximized;(4) the expression characteristic of4CL1gene was similar with that of PALI in both environments. In controlled condition, the expression of lodging-resistance species was significantly higher than that of lodging species in the budding and final flowering stage, and there is no significant difference between two types of lodging-resistance species under drought stress conditions;(5) The expression of the gene4CL2trended to be identical in two environments, the maximum expression is in the final flowering stage, and the expression in drought stress condition obviously higher than that in the controlled condition; the expression characteristics between two types of lodging-resistance species were similar with gene4CL1;(6) the expression characteristics of the expression of gene COMT was different in two treatments, expression curve was like parabolic in drought stress condition, while S-type in controlled environment, in the final flowering stage there was significantly higher expression in drought stress condition than controlled condition, the expression of gene COMT in lodging species is obviously higher than that with lodging-resistant species in early flowering stage and podding stage, while in drought stress treatment the result was contrary in podding stage.(7) the expression of gene CCoAOMT rapidly increased in the podding stage in both treatments, and in drought stress the expression was significantly higher than in the controlled treatment. The expression of CCoAOMT gene in lodging species was significantly higher than that in lodging-resistance species.(8) The expression characteristic of F5H doesn’t change in both environments, the expression maximum shows in the early flowering stage. The expression of lodging-resistance species was significant higher than the lodging species in the budding and final flowering stage;(9) the expression characteristic of gene CCR1was similar with that of PAL1and4CL1in both environments. In controlled treatment, the expression of lodging-resistance species was significantly higher than that of lodging species in the budding and final flowering stage. In final flowering stage the expression of lodging-resistance species was significantly higher than lodging species under drought stress;(10) in the budding stage the expression of the gene LAC4under drought stress was significantly higher than that in the controlled condition, but after the early flowering stage the expression decreased sharply in both environments, and no significant difference between them:only in the budding stage the expression of lodging-resistance species was significantly higher than that of lodging species;(11) the expression characteristic of LAC17was similar with LAC4expression under two conditions. The expression of lodging-resistance species was significantly higher than lodging plants in final flowering and podding stage under normal circumstances. But, under drought stress, the expression of lodging-resistance species was much lower than lodging species in budding, final flowering and podding stages.4) Correlation analysis gene expression and lignin traits.We had correlation analysis11genes expression with each other and those genes expression with lignin content and lignin monomer S/G, In addition, a cluster analysis of these11genes expression character was build. We use the path analysis to investigated target traits of lignin content and lignin monomer S/G with11genes expression as independent variables in each treatment condition. The main results are as follows:(1) except COMT and CCoAOMT, there were significant positive correlation and apparent synergy between most genes. COMT, CCoAOMT was no obvious correlation with other genes; even have negative correlation with PAL2and LAC4expression characters; In the drought stress treatment, there were significant positive correlations between C4H and CCoAOMT, LAC4and LAC17only significant positive correlation with PAL1and PAL2, in other genes still shows the apparent synergy.(2) Gene expression cluster analysis showed that:LAC4and LAC17expression characteristics were similar and PALI,4CL1,4CL2, CCR1and F5H genes expression were similar to each other, but COMT, C4H and CCoAOMT had their own expression characteristics.(3) The correlation coefficient analysis showed that there were significant negative correlations between LAC4and LAC17to the lignin content and lignin monomer S/G, but others genes have no obvious relationship with them;(4) The results of path analysis indicate that:PALI contribute the most direct effect to lignin content, next were4CL2and4CL1;Contrarily, LAC17expression contributes the largest negative effect to lignin content, and the next were F5H and C4H. The direct effect of these genes expression to the lignin content had changed under drought stress. CCoAOMT contribute the most direct effect to lignin content, next were4CL2and F5H; C4H expression contributes the largest negative effect to lignin content, and next were4CL1and LAC17.We clarified the effects on water content, plant height, diameter, breaking-resistant strength, lignin content and the S/G in drought stress condition; analyzed the dynamic changes of the of lignin metabolic pathways genes expression in different development stage; identified the genes with different expression genes between two conditions. All these results will be benefit for a further understanding on the regulation of lignin metabolic, the network of lignin related genes, and to increasing lignin content in stem for the enhance of lodging resistance in rapeseed. |
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