【作者】 张剑锋；

【导师】 傅廷栋； 马朝芝；

【作者基本信息】 华中农业大学 ， 作物遗传育种， 2014， 博士

【摘要】 自交不亲和性是显花植物在长期自然进化过程中形成的一种非常特殊的生殖隔离机制。自交不亲和配体和受体特异性识别、细胞内信号传导、由异花授粉向自花授粉的进化为植物学研究提供了一个经典的范例。作为一个高效的授粉可控制系统,自交不亲和性可应用于十字花科作物杂种优势利用。甘蓝型油菜自交不亲和系S-1300与亲和的保持系杂交,F1代表现为自交不亲和,可以利用保持系来繁殖自交不亲和系,但是其保持性机理尚不明确。BnARC1是一个U-box蛋白,具有E3泛素化连接酶活性,介导芸薹属中自交不亲和反应信号传导。拟南芥中是否存在BnARCl的功能同源基因参与自交不亲和反应,尚有待研究证实。本研究包括两方面的内容：第一部分是甘蓝型油菜自交不亲和保持性机理研究,第二部分是AtPUB2在拟南芥自交不亲和反应中功能分析。主要研究结果如下：1.甘蓝型油菜自交不亲和保持系筛选与鉴定筛选了甘蓝型油菜自交不亲和系S-1300的12份核心保持系,并对保持系S单倍型来源进行分析,发现所有保持系中A基因组上S单倍型来源于白菜BrS-29, C基因组上S单倍型来源于甘蓝BoS-15。2.甘蓝型油菜自交不亲和保持系遗传分析通过对S-1300与保持系后代分离群体基因型和表型鉴定分析发现,Bing409中保持性的遗传受单基因控制,Bing409与S-1300中A基因组上S位点的差异决定了亲和性表型,且S-1300中的S单倍型对Bing409的为显性。89008中保持性不由单基因控制,除S位点以外,还存在其他基因影响自交不亲和性表型。3.甘蓝型油菜保持性机理分析基于甘蓝型油菜自交不亲和保持系S位点基因表达分析结果,对保持性机理进行分析。在保持系中,当SPll基因不表达,自交不亲和特异性识别反应无法正常进行,因此不能抑制自身花粉表现为自交亲和；当S位点基因正常表达时,可能存在S位点以外基因影响自交不亲和反应使保持系表现为自交亲和。在S-1300与保持系的F1代中,花粉中BrSP11-29的同源基因均不表达,有功能的BrSP11-60的同源基因能够正常表达,从而使得F1表现为自交不亲和。4. AtPUB2在拟南芥中自交不亲和反应中的功能分析敲除AtPUB2后,携带有自交不亲和基因AlSRKBb-SCRb的拟南芥Col-0仍然表现为自交不亲和。拟南芥中AtPUB2的ARM-repeats结构域不与SRKb的胞内激酶结构域发生直接相互作用。AtPUB2不在自交不亲和反应发生的柱头乳突细胞中高表达,而主要在柱头乳突细胞以下的基本组织和维管组织中表达。AtPUB2不是BnARC1在拟南芥中的同源功能基因,不参与拟南芥中自交不亲和反应。更多还原

【Abstract】 Self-incompatibility is a special reproduction barrier and adopted by many flowering plants in evolution. The self-incompatible ligand-receptor specific recognition, intracellular signal transduction and the evolution from cross-pollination to self-pollination were studied as a classic example in botany research. As an efficient pollination control system, self-incompatibility can be applied in cruciferae heterosis utilization. The maintainers can be used to breed self-incompatible lines because the F1are still self-incomatible when they crossed with self-incompatible line S-1300(Brassica napus). But the genetic mechanism is not clear. BnARC1is a U-box protein with E3ubiquitin ligase activity, mediated self-incompatibility signal transduction response in Brassica. Whether there is a functional ortholog of BnARCl that assumed the role of BnARC1in self-incompatible pollination response in Arabidopsis thaliana still need to be confirmed.This study consists of two parts of work. The first part is molecular mechanism of recessive self-compatibility in Brassica napus. The second part is functional analysis of AtPUB2in self-incompatible pollination response in A. thaliana. The main results are outlined as follows:1. Screening and identification of self-incompatibility maintainers in B. napus12core maintainers of self-incompatible line S-1300were screened. The S haplotype analysis showed that the S haplotypes were derived from BrS-29on A genome and BoS-15on C genome in all maintainers.2. Genetic analysis of recessive self-compatibility in B. napusAnalysis of S locus genotypes and phenotypes in S-1300X maintainers segregation populations revealed that the recessive self-compatibility of Bing409was controlled by a single locus. The phenotype was decided by A genome S locus genotype. And the self-incompatibility of S-1300is dominant to Bing409. The recessive self-compatibility of89008was not controlled by a single locus. There must be other genes besides S locus control the SI phenotype.3. Analysis of recessive self-incompatibility mechanism in Brassica napusAccording to the expressing result of S locus genes, analysis of recessive self- compatibility molecular mechanism was performed. The maintainers were self-compatible. Because the specific recognition was inhibited when the SP11gene was not expressed, or there must be some other genes besides S locus affect the SI response when the SP111gene expression was normal. The F1from S-1300×maintainers were self-incompatible. Because the BrSP11-29ortholog was not expressed but the functional BrSP11-60ortholog expression was normal.4. Functional analysis of AtPUB2in self-incompatible pollination response in A. thalianaThe self-incompatibility response was neither abolished nor weakened by knocking out of AtPUB2. The AtPUB2ARM-repeats domain was not interacting with the SRKb kinase domain. AtPUB2was predominantly expressed in the cells of ground tissue and vasculature below the stigma epidermis but not in the sigma papilla cells. AtPUB2is not a functional ortholog of BnARCl and is not involved in SI response.更多还原