【作者】 张凤兰；

【导师】 郝丽珍； 张鲁刚；

【作者基本信息】 内蒙古农业大学 ， 蔬菜学， 2009， 博士

【摘要】 沙芥属植物是集药用、饲用、保健、防风固沙为一体的多功能沙生蔬菜,属于十字花科二年生草本植物,是蒙古高原沙地特有属,包括沙芥和斧翅沙芥(斧形类型、宽翅类型、距果类型和齿冠类型)两个种,其中沙芥为中国的特有种。本论文通过研究沙芥属植物叶片的营养成分、子叶叶表皮超微结构、遗传多样性及亲缘关系的SSR和SRAP分析以及沙芥属与十字花科部分属植物亲缘关系的SRAP分析,以利揭示属内种间、居群间和居群内多态性的差异,为该属种间的分类及其抗逆机理研究提供形态学依据,为沙芥种质资源评价和保护提供科学依据。其主要研究结果如下:1.在种质资源的考察和搜集中获得了4份新的种质,丰富了沙芥属的种质资源。2.沙芥和斧翅沙芥叶片在维生素、氨基酸等营养成分含量上存在较大差异。3.采用扫描电镜观察和定量描述了沙芥和斧翅沙芥(斧形类型、宽翅类型和距果类型)的子叶表皮的微形态特征,并发现斧翅沙芥也同沙芥具有禾本科所特有的旱生结构特征——泡状细胞,且沙芥属植物在子叶叶表皮上存在丰富的多样性。4.确定了适于沙芥属DNA提取的方法——改良CTAB法,,并建立、优化了沙芥种SSR和SRAP反应体系(10μL)。(1) SSR反应体系:Mg2+、dNTPs和引物的最适浓度分别为1.5mmol/L、0.2 mmol/L、0.2μmol/L,Taq酶0.2U,模板DNA 20～30 ng。(2) SRAP反应体系:Mg2+、dNTPs和引物的最适浓度分别为1.5mmol/L、0.2 mmol/L、0.2μmol/L,Taq酶0.3U,模板DNA 70ng,两种标记体系均稳定可靠,且这两种标记的反应体系均可用于斧翅沙芥种。5.油菜EST-SSR引物在沙芥属中的通用性研究结果表明:EST-SSR引物在沙芥和斧翅沙芥中的扩增转移成功率分别为1.96%,1.5%。6.沙芥7个居群遗传多样性SSR和SRAP分析(1)沙芥7个居群遗传多样性SSR分析在沙芥7个居群中,10对引物共扩增出40位点,多态性位点百分率为90%,多态性位点比率为40.00%～62.50%;Nei’s基因多样性变化范围为0.1230～0.2109,Shannon多态指数的变化范围为0.1872～0.3162;沙芥种居群遗传分化71.63%存在于居群内,28.36 %存在于居群间。(2)沙芥7个居群遗传多样性SRAP分析以14对引物对沙芥7个居群进行遗传多样性SRAP分析的结果表明: 14对引物共扩增出138个位点,多态性位点百分率为92.7%,居群多态性位点比率变化范围为56.20%～71.53%;Nei’s基因多样性变化范围为0.2036～0.2338,Shannon多样性指数的变化范围0.2999～0.3589;沙芥居群的遗传分化70.82%存在于居群内,29.18%存在于居群间。SSR和SRAP对沙芥7个居群遗传多样性分析的结果均表明,沙芥居群的遗传多样水平较低。7.沙芥属植物遗传多样性及其亲缘关系的SSR和SRAP分析(2)沙芥属植物遗传多样性及其亲缘关系的SSR分析在沙芥属植物中8对引物共扩增出45个位点,多态性位点百分率为90%,多态性位点比率变化范围为43.90%～65.85 %;Nei’s基因多样性变化范围为0.1370～0.1918,Shannon多态性指数的变化范围为0.2173～0.2957;沙芥属植物遗传分化75.18 %存在于沙芥种内和斧翅沙芥种各类型内,24.82%存在于种间和斧翅沙芥种内类型间。(2)斧翅沙芥种内不同类型的遗传多样性SRAP分析在斧翅沙芥种内不同类型中,19对引物共扩增出157位点,多态性位点百分率为90%,多态性位点比率为61.15%～76.43%;Nei’s基因多样性变化范围为0.1966～0.2513,Shannon多态性指数的变化范围为0.3017～0.33792;斧翅沙芥种遗传分化80.18%存在于类型内,19.82%存在于类型间。SSR分析均表明沙芥种的遗传多样水平要低于斧翅沙芥种,斧翅沙芥种内特殊类型的遗传多样性水平最高,其次是宽翅类型、距果类型、齿冠类型,遗传多样性水平最小的是斧形类型。8.沙芥属与十字花科部分属植物亲缘关系的SRAP分析表明沙芥属与供试的芸薹属、萝卜属、大青属、诸葛菜属、拟南芥属的亲缘关系均较远。更多还原

【Abstract】 Puionium Gaertn , biennial plants in cruciferae, including P.cornutum (L.) Gaertn.and P.dolabratum Maxim, were versatile sandy vegetable, which were also used for herbs, feeding and binding-sand, they mainly distributed in desert in Mongolian plateau. Nurtritional components of leaves, morphological characteristics of cotyledon epidermiswere, genetic diversity and phylogenetic relationship were investigated in P.Gaertn. were estimated or investtigated in P.Gaertn., and phylogenetci relationship between P.Gaertn and several genera in cruciferae was estimated. The main results showed that:1. Four new germplasms were found in the review and collected of germplasm resource, which enriched the germplames of P. Gaertn.2. There were differences in nutritional components of leaves between P.cornutum(L.)Gaertn.and P.dolabratum Maxim, for example Vitamin and amino acid.3. Morphological characteristics of cotyledon epidermis of P.Gaertn. were investigated by using scanning electron microscope (SEM), Bulliform cell, the endemic structure in Graminaceous, was firstly found in cotyledon epidermis of P.Gaertn , and there were abubandce diversity in cotyledon epidermis in P. Gaertn.4. The DNA extracted by the modified method of CTAB, can completely meet with the experiments related to SSR and SRAP marker, the reaction of SSR and SRAP of P.cornutum (L.) Gaertn. were establishee and optimized, which were steady and reliable.(1) SSR reaction system:1.5mmol/L, Mg2+, 0.2mmol/L, dNTPs, 0.4μmol/L primer, 1U Taq polymerase, 20 to 30ng template DNA were the optimal conditions in the reaction of 10μL volume;(2) SRAP reaction system:1.5 mmol/L Mg2+, 0.2 mmol/L dNTPs, 0.4μmol/L primer, 0.3 U Taq polymerase, 70 ng template DNA were the optimal conditions in the reaction of SRAP 10μL volume.5. Transferbility of Brassica.napus EST-SSR marker to P. Gaertn showed that the amplification percentage of tansferbility were relatively 1.96% and 1.5% in P.cornutum (L.) Gaertn. and P.dolabratum Maxim.6. Analysis of genetic diversity SSR and SRAP of 7 populations in P.cornutum (L.) Gaertn(1) Analysis of genetic diversity SSR of 7 populations in P.cornutum (L.) Gaertn40 loci were detected in 10 SSR primers, polymorphic loci percent were 90%; the percentage of polymorphic loci ranged from 40.00% to 62.50%; Nei’s gene diversity ranged from 0.1230 to 0.2109,shannon’s Information index ranged from 0.1872 to 0.3162 in population; There were 71.63% genetic variance within population and 28.36% among populations.(2) Analysis of genetic diversity SRAP of 7 populations in P.cornutum (L.) Gaertn138 loci were detected in 14 SRAP primers, polymorphic loci percent were 92.7 %; the percentage of polymorphic loci ranged from 56.20% to 71.53%; Nei’s gene diversity ranged from 0.2036 to 0.2338, shannon’s Information index ranged from 0.2999 to 0.3589; there was 70.82% genetic variance within population and 29.18% among populations.The results of two makers dicated genetic diversity was low in populations.7. Results of genetic diversity and phylogenetci relationship SSR and SRAP analysis in P.Gaertn showed that:(1) Results of genetic diversity and relationship SSR analysis in P.Gaertn45 loci were detected in 7 SSR primers, polymorphic loci percent were 90%; the percentage of polymorphic loci ranged from 43.90% to 65.85%; Nei’s gene diversity ranged from 0.1370 to 0.1918, shannon’s Information index ranged from 0.2173 to 0.2957,there was 75.18% genetic variance within specie or types and 24.82% among specie or type.(2)Results of genetic diversity SRAP analysis in P.dolabratum Maxim.157 loci were detected in 19 SRAP primers, polymorphic loci percent was 90%, the percentage of polymorphic loci ranged from 61.15% to 76.43%; Nei’s gene diversity ranged from 0.1966 to 0.2513,shannon’s Information index ranged from 0.3017 to 0.33792; there were 80.18% genetic variance within type and 19.82% among type.The genetic diversity were lower in P.cornutum (L.) Gaertn than P.dolabratum Maxim. The gradation of genetic diversity from high to low was special type, kuan chi type, ju guo type, chi guan type and fu xing type.8. Resuls of genetic relationship SRAP analysis between P.Gaertn and other genera in cruciferae showed that: the gentic relationship was far among P.Gaertn., B.Rapa, B.napus, B.olevalec, B.carinata, B.juncea B.nigra, Raphanus sativus.L, Isatis indigotica Orychragmus violaceus O.E.Schulz.更多还原