| As an important vegetative organ of plants,roots absorb water and nutrients from soil and provide mechanical support for plants.The development status of plant roots directly affects the growth status of plant aboveground parts,so the research on plant roots has theoretical value and practical significance.Adventitious root(AR)is an important part of plant root system,and its formation is regulated by a variety of endogenous and exogenous factors.The phytohormone auxin as an endogenous factor plays a key role in the de novo regeneration of adventitious roots in isolated leaves.Before the study,a screened early-flowering mutant was named ebss(early boiting and short silique)based on its phenotype of early bolting and short silique.Then,this study firstly carried out genetic analysis on the mutant gene,identified and cloned the mutant gene using the method of map-based cloning.Subsequently,the vector was constructed for complementation verification,and the tissue expression pattern and subcellular localization of the protein encoded by the gene were analyzed.Secondly,the phenotype of ebss was systematically described.Finally,quantitative real-time PCR(q RT-PCR)technology was used to analyze the expression of auxin-related genes eukaryotic leaves during the formation of adventitious roots,laying a foundation for further analysis of the functions of the genes in adventitious root development.In conclusion,EBSS gene not only regulates aboveground tissue development,but also plays a key role in root and adventitious root development of Arabidopsis thaliana.Preliminary analysis of EBSS gene may regulate adventitious root development by participating in auxin synthesis and signal transduction.It is of great significance to explore the development mechanism of Arabidopsis roots,especially adventitious roots.The main findings are as follows:(1)Genetic analysis showed that the ebss mutant was a recessive mutation controlled by a single gene,and the gene was located between molecular markers 1G5 and 1G6 on chromosome 1 of Arabidopsis thaliana by map-based cloning.Narrowed after positioning range,combined with the whole genome as a result,the heavy analysis found that mutations in a histone methylation transferase gene,on the 340 th base mutation,which happen on the first exon gene point mutation,bases from G to T,mutation caused by the GAA codon TAA,early termination,The coding protein changed from 1805 amino acids to112 amino acids.(2)Phenotypic analysis of ebss mutant showed that compared with the wild type,ebss mutant had multiple phenotypes such as early bloting,short filaments,small and slightly malformed anthers,short silique,large seeds with embryo death,more branches,yellow leaves,short main roots,less lateral roots,and short and thick hypocotyls.(3)The fusion of EBSS promoter with GUS reporter gene showed that EBSS was expressed in all tissues at different developmental stages of Arabidopsis thaliana.The expression of GFP labeled fusion protein showed that EBSS was localized in the nucleus.(4)The hypocotyl and euphylla of ebss were used as explants respectively,and the number of euphylla adventitious roots of EBSS was less than that of wild type;On the contrary,the number of hypocotyl adventitious roots was more than that of wild type,while the length of longest adventitious roots was shorter than that of wild type.(5)Real-time quantitative PCR analysis of adventitious root occurrence and auxin related gene expression showed that in the euphylla of the mutant cultured for 4 days in vitro,The expressions of genes LBD18 and LBD29 and auxin synthesis genes YUC2 and YUC8 in the LBD family were lower than those in the wild type.After treatment with auxin,the gene expression levels of LBD18 and LBD29,YUC2 and YUC8 were up-regulated,and the expression levels of IAA2 and IAA16,ARF7 and ARF19 were down-regulated in the mutants.These results suggest that EBSS gene may regulate the growth and development of adventitious roots in Arabidopsis thaliana leaves by affecting auxin synthesis and transduction. |
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