Functional Study Of SPR In Rice Root Development And Screening Validation Of Interaction Proteins

The roots of the plant can not only fix and support the aerial parts,absorb nutrients and water,but also synthesize many essential substances.The growth state of rice is closely related to the development of roots.However,most studies of plant development have mainly focused on the aerial part,but ignored the underground parts.The rice SPR(Short Primary Root)protein had been identified by our laboratory,which belonged to F-box family and involved in development of the primary root.The previous results had showed that once the SPR was loss of function,the primary roots became significantly shorter,while the number and length of crowns were not affected.Further studies have shown that the reason why the length of the spr mutant primary root became shorter is mainly due to the slower rate of the meristematic cell division,resulting in smaller meristems.In order to further analyze the mechanism of SPR how to regulate rice root development,the SPR protein has been chosen as the research object,and a series of genetics,molecular biology,biochemistry experiments have been carried out to analyze its molecular function.The main results are as follows:1.To further confirm the function of SPR gene in rice root development,the SPR complementary verification(SPRpro:SPR/spr)have been done.Five T0 positive transgenic individuals have been obtained with the SPR expression restoring to the wild type at m RNA level.At the same time,the RNAi transgenic material(SPR RNAi)have also been created.The endogenous SPR expression of 7 T0 individuals have been decreased by more than 75%.2.The phenotype of SPR complementary verification(SPRpro: SPR/spr)and RNAi materials were observed.The length of primary root of the complementary material(SPRpro:SPR/spr)have recovered to the wild type,while the length of primary root of the RNAi material is between wild type and the mutant(spr).The cytology sections of root tip have showed that the recovery of SPR gene expression could recover the length of the apical meristem cells,thereby recover the length of the primary root.These genetical and cytological results have indicated that the SPR gene does regulate the development of the primary root of rice.3.The OsSEU3 protein interacting with SPR has been successfully screened by yeast two-hybrid assay.Besides,the Bi FC experiment in tobacco has also showed the interaction between SPR and OsSEU3.The analysis of protein structure has indicated that the OsSEU3 protein belongs to the Ldb family and is the homologous gene of At SEU.The OsSEU3 gene was expressed highest in leaves and seedlings,higher in roots,endosperms and ears,and expressed less in stems and stamens.The results of q RT-PCR has showed that the transcription of OsSEU3 has been mainly concentrated on the meristematic zone of roots,which was similar to the expression pattern of SPR in roots.4.It has been proved that the OsSEU3 is localized in the nucleus.Protoplast transcriptional activity analysis revealed that the OsSEU3 gene is a transcriptional activator and can be induced by exogenous gibberellin,abscisic acid,and jasmonic acid.In summary,we have identified the OsSEU3 protein which is interacted with SPR protein.They may work together in the apical meristem of root,regulate the development of the primary root through affecting the primary root length of the meristem.