Cloning,Expression And Function Analysis Of Wheat TaTHOC1 Gene

Stripe rust,caused by Puccinia striiformis f.sp.tritici(Pst),is a destructive disease of wheat worldwide.The large variety of races and strong variability of stripe rust pathogen is among the most important constraints to global wheat production and quality.The identification and characterization of wheat disease resistance relevant genes can provide a reference for the research of wheat disease resistance mechanism and genetic breeding.THO is a kind of important protein in plant THO/TREX complex protein family.Existed studies in Arabidops is showed that it widely participated in regulating plant growth and abiotic stress.Previously,we discovered a possible candidate gene related Yr41 against stripe rust pathogen in wheat CN19.To obtained its sequence,we chose a reference sequence BE604911,which is close to the Yr41 gene in the fine mapping map(0.06 cM)and annotated to be a member of THO family.In order to obtain this candidate gene of Yr41 and study whether THO family genes play a role in wheat stress resistance and growth,we cloned TaTHOC1 gene in wheat CN19 and My11.The expression pattern of this gene was analyzed and its function was preliminarily analyzed.The main results are as follows:1.TaTHOC1 gene was cloned from wheat for the first time,based on the reference sequence obtained from the blast between BE604911 EST sequence and wheat genome database.The open reading frame of TaTHOC1 contains 1917 bp,encoding 638 amino acids,and containing an efthoc1 conservative domain.It has high homology(93%)with THOC1 gene both in Aegilops tauschii and Brachypodium distachyum.2.The tissue-specific expression pattern of TaTHOC1 gene showed tissue-specific profile.The expression level of this gene in grains and leaves was significantly higher than that of other tissues(P<0.01): grains > leaves >flowers > glumes > stems > roots.As for the treatment of cold stress and salt stress,the expression of TaTHOC1 gene in wheat was both decreased,with no significance.Moreover,after the treatment of osmotic stress,the expression of TaTHOC1 gene was firstly increased and then decreased.3.Analys is of the expression level of TaTHOC1 gene in wheat infected by CYR32 and V26(CYR34)showed that there were differences in the expression patterns of TaTHOC1 in compatible/ incompatible wheat.And the expression patterns of TaTHOC1 were also different after being infected by different physiological races.After infection by CYR32,the expression of TaTHOC1 gene in affinity interaction(CYR32/My11)is gradually increased.In incompatible wheat CN19,the expression of TaTHOC1 gene generally showed a downward trend.During the infection of physiological raceV26,the expression pattern of My11 is similar to that of CN19.In affinity interaction(V26/My11),the expression of TaTHOC1 gene firstly increased and then decreased.In the non-affinity interaction(V26/CN19),the expression of TaTHOC1 gene was up-regulated.The immune response triggered by Chitin,the molecule related to fungal pathogens,was simulated in wheat plants and cells.The results showed that the expression of TaTHOC1 gene was sharply down-regulated and showed different expression patterns from TaCMPG1,TaPDR2 and TaPR1.These genes are all reporter genes in wheat immunity.Therefore,it showed that the contribution of TaTHOC1 gene to plant resistance might be realized via a negative way.4.The expression profiles of TaTHOC1 was characterized in CN19 and My11 treated by 4 kinds of exogenous hormones(ABA/SA/J A/ET).The results showed that after different hormone treatments,the response of TaTHOC1 was different,and its expression characteristics were different among varieties.In disease-resistant wheat CN19,both hormones ET and ABA can rapidly induce the up-regulated expression of this gene.While after being treated with hormones SA and MeJA,this gene is rapidly down-regulated and then up-regulated.However,in susceptible wheat My11,the expression pattern of TaTHOC1 gene response hormone is opposite to that of resistant wheat.5.The roles played by TaTHOC1 gene in GA-mediated wheat grain development were also discussed.The results showed that after GA treatment of wheat grain,TaTHOC1 gene of resistant/ susceptible wheat all responded to GA,and the expression trend of TaTHOC1 gene was consistent with that of GA responsive gene TaGASA4.However,the expression pattern of TaTHOC1 gene in the process of wheat grain development is different from that of TaGASA4.As the grain matures,the expression of TaTHOC1 gene gradually decreases,but it is not significant.However,the expression of TaGASA4 gene was significantly increased and then gradually decreased,suggesting that TaTHOC1 gene might not participate in GA-mediated grain development process.6.Subcellular localization of p TaTHOC1 was carried out by transient transformation in wheat protoplast,and the results showed that TaTHOC1 was located in the nucleus of wheat cell.Meanwhile,the overexpression vector of this gene was constructed,which laid a foundation for subsequent transgenic experiments to verify the gene function.