Screening Of T-DNA Insertion Auxin Related Mutants In Brachypodium Distachyon L. And The Role Of BdWAR1 In Interaction Between Auxin And ABA

Brachypodium distachyon is a new model plant for crops.Compared to the traditional model plant Arabidopsis thaliana and rice,it has many advantages.Auxin is an important phytohormone.It is involved in various processes of plant development,such as maintenance of apical stationary center,the differentiation of root tip cells,phototropism and geotropism.Auxin regulates transcription by ubiquitin(Ub)-mediated degradation of Aux/IAA repressors.Auxin binds to the auxin receptor SCF~TIR1/AFBsIR1/AFBs and promotes AUX/IAAs degradation,releasing ARFs and induces auxin response.ABA is an important phytohormone a small molecule that function to inhibit growth and to regulate plant stress responses.ABA binds ABA receptors PYR/PYLs and inhibits PP2Cs'activity,and promotes SnRK2s and its downstream transcription factors ABI3 and ABI5's function finally induces ABA response.Currently,the crosstalk of auxin and ABA signaling pathway has become a hot research topic in developmental biology.In this study,Brachypodium distachyon were taken as the research object,we made auxin response screening of T-DNA insertion mutant library.There are my results:(1)The concentration of NAA was determined to be 1000nM and the concentration of ABA was determined to be 5?M.(2)Screening the T-DNA insertion mutant library,we found nine mutants with changed auxin response.Four of them had been located the T-DNA insertion site,such as bdwar1,NO.35.There were six short root mutants,such as bdwar1,NO.12,NO.31.Besides,There was an early flowering mutant NO.7.In addition,there was an auxin sensitive mutant NO.18,and two auxin insensitive mutants,NO.12 and NO.31.(3)The bdwar1 was found to be short root,insensitive to NAA,hypersensitivity to ABA and stress,slim seeds,fewer branches,short and narrow leaves and large leaf inclination,long and more root hair on the new maturation zone.(4)In bdwar1,we located the T-DNA insertion site at-262bp before ATG of BdWAR1.The result of qRT-PCR revealed that the relative expression level of BdWAR1 was dipped to 60%.(5)Through the comparison of amino acid sequences,we found there were four homologous genes in Arabidopsis thaliana.And wheat,maize and rice have one homologous gene,respectively.And we found two or three conserved domain,which may be the position signal or active site.(6)Observed expression pattern,we found that BdWAR1 tend to express in vascular bundle.(7)The qRT-PCR results revealed that there are many changes of relative expression level in bdwar1.Most of them were related to Auxin pathway,ABA pathway and root hair developmental pathway.(8)Yeast two hybrid experiments showed that the BdWAR1 interacted with ABI1 and TPL.