Cloning And Functional Study Of DROs Frommaize And Mexico Teosinte

As an important food crop,economic crop and energy fuel in China,the demand for corn has gradually increased in recent years.However,due to frequent h?Man activities and intensified global climate change,freshwater resources are in short supply in many regions of the world,and the area of arid regions is expanding day by day.Therefore,improving the ability of maize to cope with drought plays an important role in increasing maize yield in arid areas.OsDRO1?DEEPER ROOTING 1?gene is a gene found in rice that affects root distribution by controlling the included angle of root growth,thus affecting drought resistance of plants.Through amino acid sequence alignment,we found that there may be two homologous genes of OsDRO1 in maize,named ZmDRO1 and ZmDRO2 respectively,both of which are located at the proximal end of the long arm of chromosome 7.Teosinte is the natural ancestor of maize and has good performance in resisting abiotic stress.The root system of Zea mays ssp.Mex.icana?Abbreviated as Mex.?is quite different from that of cultivated maize B73.Omori used Zea mays ssp.luxurians and cultivated maize B73 to construct a recombinant inbred line and then targeting 10 QTLs for root angles control.Among them,4 major QTLs are concentrated in the proximal end of the long arm of chromosome 7,including ZmDRO1 and immediately adjacent to ZmDRO2.In order to further explore the contribution of DROs to the root system differences of Mex.and B73 and its role in drought avoidance,the genes of DRO1and DRO2 of Mex.were cloned and compared with those ZmDRO1 and ZmDRO2.Meanwhile,the function of DROs gene in maize was studied by physiological,biochemical and molecular biological methods.The specific results are as follows:1.The similarity of amino acids encoded by DRO1 gene of Mex.and B73 is as high as98.4%,and the difference of gene sequence is mainly concentrated in the part before exon 3;The amino acid similarity of DRO2 gene is 97.3%,and the differencr of gene sequence is mainly concentrated in the promoter region.2.DRO1/2 gene responds to ABA induction.The expression trend of DRO1 gene in B73and Mex.was first rising and then callback.The background expression of DRO1 gene in Mex.is lower than that in B73,but the up-regulation multiple is higher than that in B73 after ABA treatment.3.Subcellular localization results showed that ZmDRO1 and ZmDRO2 were mainly located in cell membrane and a small amount in cytoplasm,and the localization results of tobacco and maize protoplasts were consistent.4.After drought treatment,the distribution ratio of Mex.root system in deeper soil layer is higher than B73,and the reduction ratio of dry weight of above ground part of Mex.is significantly lower than B73.Parental separation population constructed by B73 and Mex.was genotyped at BC₂F₈,and DRO1 materials with different background sources were selected to measure drought resistance,root included angle and root distribution.The results showed that the biomass reduction rate of DRO1 genotype Mex.material was significantly lower than that of B73?P<0.05?.Under non-drought stress,the angle between DRO1 genotype and Mex.material is significantly smaller?P<0.05?than that between DRO1 genotype and B73 material.5.DRO1/2 was driven by ABA-induced promoter to express,and Arabidopsis thaliana and maize materials with high DRO1/2 expression under ABA-induced conditions were obtained.It was expected that DRO1/2 could be highly expressed under mild drought stress,so as to improve drought tolerance of transgenic plants.6.Because DRO1 is highly expressed in coleoptiles,it is speculated that it may be related to light signal response,and the expression of DRO1 gene was detected under light and dark conditions.The results showed that light could inhibit DRO1 gene expression at transcription level.In order to further analyze the possible functions of DRO1 in light signal transduction,we selected light receptors PHOT1,PHOT2 and PHYA,and using BiFC to detected whether there is interaction between them and DRO1.The results showed that the proteins of ZmDRO1interacted with the proteins of ZmPHOT1,ZmPHOT2 and ZmPHYA on the cell membrane in pairs.