The Study On The Function Of Auxin Biosynthetic Gene-OsYUCCA2 In Rice

Auxin is an important phytohormon in regulating plant growth and development.Os YUCCA2(Os YUC2)is one member of the auxin biosynthesis key gene family Os YUCCAs in rice.The function and development and tissue specific expression of Os YUC2 gene was studied using rice(Oryza sativa L.japonica.CV.Nipponbare).We analysed the effects of different treatments on the expression of Os YUC2 gene at the level of transcription by using semi-quantitative RT-PCR.The results showed that the expressive level of Os YUC2 gene was higher in the DMTU(H2O2 scavenger)and GSH(glutathione)-treated roots but was higher in the Cd(NO3)2 and BSO(glutathione synthesis inhibitors)-treated leaves than in the control roots/leaves.The results suggest that the Os YUC2 gene differentially expressed in roots and leaves.To study the function of Os YUC2 gene,the p Ubi-Os YUC2-GUS forward and reverse fusion expression vectors were constructed respectively.The results as follows:(1)When compared with the wild type rice the growth and differentiation of the resistant callus was slower in PUbi-Os YUC2-GUS reverse expression transgenic rice.Only a few differentiated seedlings were obtained,and these seedlings died rapidly during the experimental periods.The results indicated that the reverse expression of Os YUC2 gene inhibited the growth and development of rice.(2)The growth and differentiation of the resistant callus was faster in PUbi-OsYUC2-GUS forward expression transgenic rice as compared to that of the wild type rice.During the experimental periods 120 transgenic plants were obtained.(3)Phenotypes of some of the transgenic rice were analyzed.Compared with the wild type rice seedlings,the length and number of adventitious roots,the plant shoots and the width of leaves increased,but the main vein was thinner in PUbi-Os YUC2-GUS forward expression transgenic rice.These results indicated the forward expression of Os YUC2 promoted the growth and development of rice in a certain extent.(4)The resistance of PUbi-OsYUC2-GUS forward expression transgenic rice to copper stress was studied by treating seedlings with Cu SO4.The results showed that the water loss declined in seedlings,the accumulation of H2O2 decreased in adventitious roots,the content of MDA reduced in root system and shoots,and the relative water content was more in transgenic rice than in the wild type rice seedlings.The results implied that the forward expression of Os YUC2 increased the ability of rice plants in resisting copper stress to some extent.(5)The development and tissue specific expression of Os YUC2 was analyzed by detecting GUS activity.The results indicated that the GUS activity was higher in the plantule,young roots,young leaf and root tips at the early stages of growth and development.However,the GUS activity decreased gradually until disappeared with the growth of leaves and roots.In the transverse section of the root tips,the GUS activity gradually decreased from the meristematic zone to the maturation zone.The distribution of GUS staining was observed in all cells in the meristematic zone but only in part cells in maturation zone.In the transverse section of the leaf tips,the GUS staining mainly distributed in mesophyll cells and vascular bundles.To further study the tissue and organ specific expression of Os YUC2 the POs YUC2-EGFP fusion expression vector was also constructed and transformed into rice.When the POs YUC2-EGFP transgenic rice seedlings were treated with PEG6000 for 8 h and Cu SO4 for24 h,the GFP signal was observed in the meristematic zone of adventitious and lateral roots.Furthermore,the distribution of GUS staining in the PUbi-Os YUC2-GUS forward expression transgenic rice was similar to that of the DR5-GUS transgenic rice.The activity area of GFP in POs YUC2-EGFP transgenic rice was overlapped with the active region of GUS in the root tips of DR5-GUS transgenic rice.These results implied that the auxin biosynthesis controlled by Os YUC2 gene was a part of the total auxin.Taken together,these results showed that the expression of Os YUC2 was mainly in the splinter cells,and its expression was specific in early developmental stages,in some tissues and organs.Thus Os YUC2 played important regulatory roles in the early stages of growth and development in rice.