Morphology And Molecular Basis Of Chrysanthemum Root System Responding To The Nitrate Nitrogen

Chrysanthemum is one of the four major cut flowers in the world.Its'cultivation areas and yields are in the front rank of flowers.As we all known,chrysanthemum is fertilizer-sensitive plant,nitrogen?N?is essential for the growth and development of chrysanthemum.In well ventilated soils,NO₃^--N is the main inorganic N source for root absorption,and the absorption of NO₃^-is mostly achieved by the nitrate transporters?NRTs?in the root of chrysanthemum.NO₃^-is not only the main N source of plant nutrition,but also acts as a signal molecule to regulate the expression of related genes to affect the growth and development of plant root.Therefore,the rooted cuttings of chrysanthemum were treated by Hogland nutrient solution with 10 mmol·L^-1KNO₃ while control was treated without nitrogen.The root morphology,anatomical structure,the contents of NO₃^-,photosynthetic characteristics,the contents of IAA and CTK in the roots and leaves were measured,and the relative expression levels of CmNRTs and CmANR1 were also measured using quantitative real-time PCR after treatment.The aim of our experiment is to reveal the morphological structure and molecular basis of chrysanthemum root system response to NO₃^-signal,and to provide a theoretical basis for the selection and breeding of new varieties of chrysanthemum efficiently utilized.The results are as follows.1.After treated with 10 mmol·L^-11 KNO₃ while control was treated without N.The roots were selected to determine their morphology parameters on the 0,1,3,7,14,21,and 28 day.The results showed that:the overall length,average diameter,total surface area and total volume of roots were increased significantly after 7 days compared with those of the control.As the longer processing time,the greater increased.At 28th,the root morphology indicators of NO₃^-treated chrysanthemum increased by 203.63%,59.80%,129.33%,114.33%,and105.04%,compared with the control,respectively.2.The cross section of chrysanthemum roots on the 28th day was observed with a scaled biological microscope,and the vascular bundle size of each side of the roots were determined.The results showed that the diameter of the vascular bundle of the NO₃^-treated chrysanthemum primary roots were 0.103 mm,the vascular bundle diameter of 2nd root was about 0.107 mm,and the vascular bundle diameter of 3rd root was about 0.102 mm.Compared with the control,the ratio of vascular bundle diameter and ratio of vascular bundle to root cross section of the 1st,2nd,and 3rd roots after treatment had increased to varying degrees.The vascular bundle diameters of the 1st,2nd,and 3rd roots increased by 5.10%,13.83%,and 5.15%,respectively.The ratio of vascular bundles to the root cross-sectional diameter increased by 15.70%,13.80%,and 12.42%,respectively.3.The nitrate nitrogen contents in roots and leaves were measured on the 0,1,3,7,14,21,and 28 day.The NO₃^-contents reached the peak on the 7th?0.45 mg·g^-11 FW?in roots and on the 14th?0.35 mg·g^-11 FW?in leaves,respectively.Then both of them decreased slightly,but they were maintained higher level compared with those of the control all days.However,the nitrate nitrogen content was always at a relatively high level compared with those of the control?controls'nitrate nitrogen contents of root and leaf were maintained at 0.17-0.27mg·g^-11 FW and 0.16-0.22 mg·g^-11 FW,respectively?.4.The net photosynthetic rate?P_n?,chlorophyll fluorescence parameters(F_o,NPQ and F_v/F_m,?_PSII)and chlorophyll content were determined on the 0,1,3,7,14,21,and 28 day.The results showed that compared with the control,at 28th,the P_n at the leaves of chrysanthemum rose by 29.4%after treatment with NO₃^-.The F_o and NPQ decreased by24.4%and 22.7%,respectively.And the F_v/F_m and?_PSIISII increased by 7.3%and 36.1%,chlorophyll contents also increased by 29.4%.All of these indicated that the photosynthetic characteristics of NO₃^-treated leaves were better than those of the control,and the effect of NO₃^-treatment on the photosynthetic characteristics of chrysanthemum showed a significant time effect.5.The contents of IAA and CTK in roots and leaves were determined on the 0,1,3,7,14,21,and 28 day.The results showed that the contents of IAA and CTK in NO₃^-treated roots and leaves both increased significantly compared with the control.The peaks of IAA and CTK contents in roots appeared on the 7th day,the peaks in leaves appeared on the 14th day.The IAA and CTK contents of control did not change significantly.Although the trends are similar,there is significant difference in IAA and CTK content in leaves and roots.6.The expression levels of CmNRT1.1,CmNRT2.1,CmNAR2.1 and CmANR1 were determined on the 0,1,3,7,14,21,and 28 day.After NO₃^-treatment,the peak of relative expression level of CmNRT1.1,a double-affinity nitrate transporter gene,appeared on the first day.The relative expression level of CmNRT2.1 and CmNAR2.1,peaked on the third day.And the relative expression level of CmANR1,a specific gene for lateral root development of chrysanthemum reached the peak on the 7th day.After NO₃^-treatment,the relative expression levels of these 4 genes were always at a relatively high level compared with those of the control.And the expression trends were also similar,they all increased first and then decreased,and remained relatively stable.This indicates that their expression is induced by NO₃^-signal,but the relative expression peaks are different and the degree of induction is also different.The internal and external environmental signals of plants jointly affect the growth and development of the root system.NO₃^-treatment changed the morphological structure of the root system of chrysanthemum and promoted the growth of lateral roots.In this process,the root vascular bundle structure,nitrate nitrogen contents,photosynthetic characteristics,endogenous hormone levels and related gene expression levels have also changed.The results indicated that the root system of chrysanthemum could respond to the NO₃^-signal in the growth medium by up-regulating the expression of CmNRTs and CmANR1 genes.On the other hand,suitable NO₃^-increases the contents of IAA and CTK in the root,and both of them improve the root system.They can increase the absorption and utilization of NO₃^-in the root,and promote the growth and development of chrysanthemum.