【作者】 刘宇；

【导师】 刘志斌；

【作者基本信息】 四川大学 ， 林业硕士（专业学位）， 2023， 硕士

【摘要】 干旱影响自然界植物分布,限制农业生产,威胁粮食安全。小野荞麦是蓼科荞麦属植物,该属还包括重要经济作物苦荞和甜荞。苦荞因含有丰富的类黄酮物质而广受欢迎,但是,苦荞主产区多为干旱半干旱地区,干旱是限制其产量及品质的主要环境因素。为了培育苦荞耐旱品种,找到荞麦属内的耐旱野生植物资源,我们开展了该属植物的野外调查和采集,并从小野荞麦中挖掘到一个与耐旱相关的基因,初步研究了其功能。荞麦属野外调查发现小野荞麦生境恶劣,为极度干旱瘠薄的荒山坡、碎石堆、岩石缝等,种群数量少,植株矮小,整株呈红褐色,花多,花结构具有较典型的荞麦属植物特征。将小野荞麦在实验室驯化栽培,与野外生长的植株相比,其形态发生显著变化,株高明显增加,整体植株呈绿色,但人为干旱后,植株颜色由绿色转变成近似于野外观察到的红褐色。查阅苦荞抗逆相关文献,并通过同源克隆方法从小野荞麦中得到一个耐旱相关基因FlbZIP12,发现其定位于细胞核,而且在根中表达量最高,受到ABA、PEG和NaCl的诱导表达。Y2H和BiFC结果显示FlbZIP12与ABA信号通路关键调控因子FlSnRK2.2和FlSnRK2.6具有相互作用。PEG处理后,过量表达FlbZIP12的苦荞毛状根中SOD和CAT活性增加而MDA含量下降,抗逆响应基因FtRD29B和FtERD1都上调表达。类黄酮合成途径关键酶基因FtC4H的表达量在3个过表达FlbZIP12的苦荞毛状根株系中都极显著增加,FtF3H、FtF3’H、FtFLS、FtPAL、Ft4CL和FtCHS在3个株系中的表达量不完全一致,在OE#5株系中,这7个基因都极显著上调表达。这些结果说明FlbZIP12通过与FlSnRK2.2和FlSnRK2.6相互作用参与到ABA信号通路以提升苦荞毛状根的耐旱性,而且FlbZIP12普遍正向调控类黄酮合成通路酶基因的表达,这可能与人为干旱胁迫后,小野荞麦植株颜色的转变有关。小野荞麦是一种优质资源植物,既可作为苦荞耐旱育种的远缘杂交亲本,又可用作生态修复先锋植物。该研究充分展示了小野荞麦的上述特征,并从中克隆了一个耐旱相关基因,也为苦荞抗旱育种提供了候选基因资源。更多还原

【Abstract】 Drought affects the distribution of plants in nature,restricts agricultural production and poses a serious threat to food security.Fagopyrum leptopodum(Diels)Hedberg belongs to the Fagopyrum Mill of Polygonaceae,which includes important cash crops Tartary buckwheat(Fagopyrum tataricum)and common buckwheat(Fagopyrum esculentum).Tartary buckwheat is popular for its abundant flavonoids.However,the biggest problem faced by Tartary buckwheat industry is low yield.However,the main producing areas of Tartary buckwheat are mostly in arid and semi-arid mountain areas.Drought is the main environmental factor that limits its yield and quality.In order to cultivate drought-tolerant varieties of Tartary buckwheat and find drought-tolerant wild plant resources in Fagopyrum,we carried out Fagopyrum Mill plants investigation and collection.Excavated a gene related to drought resistance from F.leptopodum and studied its stress resistance function.Field investigation showed that the habitat of F.leptopodum is so terrible.The population of F.leptopodum is small,and the plants are short with reddish-brown colour.What’s more,a lot of flowers had typical characteristics of Fagopyrum Mill.In the wild,we found that F.leptopodum grew in extremely dry and barren hillsides,gravel piles and rock crevices.When the F.leptopodum was domesticated and cultivated in the laboratory,compared with the plants grown in the field,its morphology changed signficantly,the plant height increased obviously,and the whole plant was green.However,after man-made drought,the plant color changed from green to reddish brown,which was similar to that observed in the field.Referring to the related literature of stress resistance of Tartary buckwheat,a drought tolerance related gene FlbZIP12 was obtained from F.leptopodum by homologous cloning method.It was found that it was located in the nucleus,and its expression was the highest in the root,and was induced by ABA,PEG and NaCl.The results of Y2H and BiFC showed that FlbZIP12 interacted with FlSnRK2.2 and FlSnRK2.6,the key regulators of ABA signal pathway.After PEG treatment,the activities of SOD and CAT in hairy roots of Tartary buckwheat overexpressing with FlbZIP12 increased,while the content of MDA decreased,and the expression of stress-resistant genes FtRD29 B and FtERD1 were up-regulated.The expression of FtC4H,a key enzyme in flavonoid biosynthesis pathway,was significantly increased in three Tartary buckwheat hairy root lines with overexpressing FlbZIP12.The expressions of FtF3H,FtF3’H,FtFLS,FtPAL,Ft4CL and FtCHS were not completely consistent in the three lines,and these seven genes were significantly up-regulated in OE#5.These results suggest that FlbZIP12 through interaction with FlSnRK2.2 and FlSnRK2.6 to participates in ABA signal pathway to improve drought tolerance of Tartary buckwheat hairy roots,and FlbZIP12 generally positively regulates the expression of enzyme genes in flavonoid synthesis pathway,which may be related to the color change of F.leptopodum after artificial drought stress.F.leptopodum is a kind of high-quality resource plant,which can be used not only as a distant hybrid parental species,but also as a pioneer plant for ecological restoration.This study fully demonstrated the above characteristics of F.leptopodum,and cloned a drought tolerance-related gene which was a candidate gene for Tartary buckwheat drought resistance breeding.更多还原