Effects Of Plant Density And 2,4-D Isooctyl Ester On Aegilops Tauschii Tillering And Its Regulatory Mechanism

Aegilops tauschii is one of the most troublesome weeds and severely infests with wheat fields in China.It widely spreads in major wheat-producing provinces of China(Shandong,Henan,Hebei,Shaanxi,Shanxi)and poses a serious threat to the yield and quality of wheat.Tillering is an important biological trait that determines the propagation coefficient of Poaceae weeds.Therefore,strong tillering ability of Ae.tauschii will lead to its high reproductivity,and then accelerate its infestation with wheat and expansion to new habitats.To understand the regulatory mechanism of Ae.tauschii tillering,the effect of plant density on Ae.tauschii tillering was evaluated.The transcriptomic,physiological and biological responses of Ae.tauschii tillering to plant density were also assayed.Meanwhile,the effect of2,4-D isooctyl ester on Ae.tauschii tillering was evaluated with Triticum aestivum as control,and the basis explaining the different tillering responses to 2,4-D isooctyl ester between the two plant species was also studied in detail.The main results are shown as follows:1.A total of 192 Ae.tauschii populations collected from the wheat fields of Chine were used to evaluate its genetic diversity and population structure with 17 SSR markers.The results showed that Ae.tauschii exhibited a moderately high level of genetic diversity,high differentiation,and limited gene flow.Compared with other studied provinces,Hubei populations possessed relatively low genetic diversity.STRUCTURE analysis suggested that Ae.tauschii populations in wheat fields of China can be divided into three groups,and this result was supported by UPGMA cluster analysis.Additionally,among the three groups,solely 7%of the total variation was detected,whereas 67%of the total variation was occurred among different populations within groups based on analysis of molecular variance.2.According to the population structure results,12 Ae.tauschii populations were selected and used to evaluate the effect of plant density on its propagation indicators,including the tiller number per plant.The results showed that plant density played a significant role in the tiller number per Ae.tauschii plant(P<0.05),and its tiller number per plant gradually decreased with the increase of plant density.Besides,the length of Ae.tauschii tiller buds located at the axil of the first leaf was assayed under different plant densities and the results showed that plant density significantly inhibited the outgrowth of its tiller bud.3.In the transcriptome analysis on Ae.tauschii tiller buds at different plant densities,differentially expressed genes(DEGs)were significantly enriched in the porphyrin and chlorophyll metabolism,photosynthesis,carbon fixation in photosynthetic organisms and plant hormone signal transduction pathways.Besides,DRMs,SPL10,FHY1,ABAox3,CDK2 genes may be involved in the regulation of Ae.tauschii tillering.The chlorophyll content and photosynthesis,soluble sugar and phytohormone levels were determined at different plant densities.The results showed that chlorophyll levels,photosynthetic efficiency and soluble sugar contents were distinctly inhibited by high plant density in Ae.tauschii.Additionally,the levels of abscisic acid(ABA)and gibberellin acid 3(GA₃)were strikingly higher at high plant density compared with those at low plant density,yet the reverse tendency was observed for indole-3-acetic acid(IAA).4.The effects of 2,4-D isooctyl ester on the tiller number per plant and tiller bud elongation in Ae.tauschii were assayed with T.aestivum as control.The results showed that 2,4-D isooctyl ester had inhibitory effects on the tiller number per plant and tiller bud outgrowth in both Ae.tauschii and T.aestivum,and this inhibitory effect was enhanced with the increase of 2,4-D isooctyl ester spraying dose.However,the tiller number per plant and tiller bud outgrowth in T.aestivum were more vulnerable to be restricted by 2,4-D isooctyl ester than that in Ae.tauschii,indicating that Ae.tauschii tillering was more insensitive to 2,4-D isooctyl ester compared with T.aestivum.5.The contact angle of 2,4-D isooctyl ester droplets on T.aestivum leaves was markedly lower than that on Ae.tauschii leaves.There were more trichomes on the Ae.tauschii leaves than on the T.aestivum leaves.During the uptake and metabolism of 2,4-D isooctyl ester,a distinct increase in 2,4-D isooctyl ester uptake was detected in T.aestivum.Besides,compared with Ae.tauschii,T.aestivum exhibited a greater accumulation of primary metabolite 2,4-D in plants.The expression level of 2,4-D isooctyl ester receptor TIR1 gene was evaluated with absolute quantitative real-time PCR(q PCR)and the results showed that T.aestivum exhibited obviously higher expression level of TIR1 gene compared with Ae.tauschii.Transcriptome analysis was performed on the tiller buds of Ae.tauschii and T.aestivum after 2,4-D isooctyl ester treatment.The results showed that DEGs in T.aestivum response to 2,4-D isooctyl ester were significantly enriched in plant hormone metabolism and signal transduction pathways,but similar change was not observed in Ae.tauschii.Additionally,a total of nine candidate genes were identified from RNA-Seq,which may be related with the different tillering responses to2,4-D isooctyl ester between Ae.tauschii and T.aestivum.Endogenous hormone levels in the basal part of stem in Ae.tauschii and T.aestivum were assayed after spray with 2,4-D isooctyl ester.The results showed that the dynamics of ABA and cytokinin(CTK)contents responded differently to 2,4-D isooctyl ester in the two plant species,suggesting that ABA and CTK participated in the 2,4-D isooctyl ester-mediated regulation of tillering.External ABA clearly restricted tiller bud growth in both T.aestivum and Ae.tauschii,while 6-benzyl aminopurine had no significant effect.Given that,the effect of plant density on Ae.tauschii tillering was clarified.The intrinsic regulatory network underlying Ae.tauschii tillering response to plant density was established based on the results of transcriptomic,physiological and biological responses.Meanwhile,with T.aestivum as control,the effect of 2,4-D isooctyl ester on Ae.tauschii tillering was determined,and the two plant species exhibited different tillering sensitivities to 2,4-D isooctyl ester.The reasons for the different tillering responses to 2,4-D isooctyl ester were clarified using the techniques of RNA-Seq,q PCR and targeted metabolomics.Such results will help us to further understand the regulatory mechanism of Ae.tauschii tillering,and provide new ideas for the establishment of control strategies against this weed species based on decreasing its tiller number per plant and propagation coefficient.