Cloning And Functional Analysis Of Wheat TaIAA8-1B Gene

In plants,auxin is sensed by its receptors,and is degraded by ligase ubiquitination,releasing the protein's inhibition of auxin response factors,thereby allowing transcription factors to bind to the downstream auxin-regulated gene response elements to regulate these gene expression transmits auxin signals.Aux/IAA protein,which acts as a class of transcriptional repressors,has been shown to play significant role in the auxin signal transduction pathway.Among them,the IAA8 gene is one of the members of the Aux/IAA family.In this study,we focus on the exploration of the biological function of TaIAA8 by molecular biology,biochemistry and physiology.The results obtained are as follows:1.We cloned the wheat TaIAA8 gene and transferred it to Arabidopsis by Agrobacterium infection.Then,we obtained the homozygous transgenic lines,the phenotype of transgenic lines showed that the main roots of the overexpression lines were not much different from that of the empty GFP lines,but the number of lateral roots were less.These results suggested that overexpression of TaIAA8-1B may promote the response of Arabidopsis roots to auxin.2.We examined the expression patterns of TaIAA8-1B genes in different tissues during different periods.RT-PCR analysis showed that TaIAA8-1B was highly expressed in the leaves at seedling stage.However,compared with other tissues,the expression level in roots was higher at heading stage and jointing stage.3.The expression pattern of wheat TaIAA8-1B in roots also were evaluated under abiotic stress conditions.The results showed that the expression of TaIAA8-1B had not change significantly under PEG,Heat and 4?,indicating that the TaIAA8-1B gene would not be affected under these conditions.4.Construction of pCAMBIA1300-TaIAA8-1B-GFP recombinant vector for transient expression,detection of fluorescent signal,and observation of the subcellular localization of TaIAA8-1B.The results showed that TaIAA8-1B was mainly located in the nucleus5.Through the screening of the yeast library,the possible interaction protein ARF21 was obtained.The yeast two-hybrid experiment was used to verify the protein interaction.As a result,the IAA8 protein interacted with the ARF21 protein.6.Through the association analysis of the wheat gene TaIAA8-1B mutation site(C/T)and agronomic traits revealed that the allelic mutation C was significantly related to the deeper root depth of wheat,and was positively selected in the history ofwheat breeding.From the above experiments indicated that TaIAA8 was mainly located in the nucleus,the highest expression level in wheat seedling stage leaves,and its expression was not affected by abiotic stress such as PEG.Further research suggested that TaIAA8 responds to auxin signals and interacts with TaARF21 to regulate lateral root growth.