【作者】 张睿鹂；

【导师】 张启翔；

【作者基本信息】 北京林业大学 ， 园林植物与观赏园艺， 2008， 博士

【摘要】 报春花属（Primula）,世界著名的高山花卉,以其丰富绚丽的花色、各具特色的花型而备受世界花卉园艺爱好界的关注和喜爱。中国西南横断山区是报春花属现代的分布中心和多样化中心,其中云南有报春花属120余种。但是继20世纪90年代初期中国报春花科植物志的编撰工作完成之后,由于受到环境因素和人为因素（基础设施修建和旅游开发等）的影响,报春花属植物资源开始遭到破坏;同时由于报春花属植物多分布在山地高海拔区域,夏季不耐高温的习性严重限制其在中国的引种驯化和园艺开发进程。因此本研究以植物志、相关文献和标本为线索,采用线路法结合样地对滇西北地区报春种质资源进行调查,以求了解报春花具体的分布情况和种群的生存状态等;同时选取其中观赏性状突出、具有较大引种驯化的报春进行群体的遗传分析,以制定合理的保护策略。在引种驯化方面,通过相关技术措施的研究以提高报春的耐热性,同时采用mRNA差别显示技术研究报春对热激反应的差异性表达。主要结论如下:1.历时3年在滇西北地区共计调查到报春55种（含亚种和变种）,分布在18个组。本地区的报春种类占云南的近50%,并且组的分布占云南分布组的95%以上,本地区的报春种质资源丰富。种质资源具有物种多样性丰富,种群密度变化大;分布呈现明显的垂直性和集中性;生境类型复杂多样等特点。但是由于受到人为因素和自然因素的影响,报春种群的分布面积和分布范围正在减少,其中以苍山分布的薄叶粉株报春（P. membrannifolia）和丽江分布的小苞报春（P. bracteata）这两种报春的破坏程度最严重。其余多数种报春分布在海拔3000m以下的种群都受到严重的破坏,致使滇西北报春集中分布的区域由海拔2000～4000m缩减为海拔3000～4000m。2.根据观赏性状和生态因子的综合量化评价,铁梗报春（P. sinolisteri）、巴塘报春（P. bathangensis）、多脉报春（P. polyneara）、灰岩皱叶报春（P. forrestii）、海仙报春（P. poissonii）、香海仙报春（P. wilsonii）、中甸灯台报春（P. chungensis）、钟花报春（P. sikkimensis）、峨嵋报春（P. faberi）、紫花雪山报春（P. sinopurpurea）、中甸海水仙（P. monticola）、滇北球花报春（P. denticulate ssp.sinodenticulata）、高穗报春（P. vialii）、羽叶穗花报春（P. pinnatifida）和穗花报春（P. deflexa）具有较高的引种驯化价值。3.滇北球花报春野外共计调查到10个群体,分布在大理、漾濞、云龙、丽江、维西等9个区域,其中植物志和标本记载有分布的昆明、嵩明、易门、玉溪、丽江暂着山和泸水没有找到分布群体。与标本记录相比,群体的分布范围和分布面积都有一定程度的减少。构建10个群体初级核心种质300份。通过表型变异和遗传多样性性评价,其中有5个群体参与核心种质的构建;通过捕获曲线模拟遗传多态性,每个群体随机抽取27～31株,即可捕获95%以上的群体遗传多态性。滇北球花报春的核心种质包括种子、种苗和植物DNA三种形式。4. Ca²⁺参与作为植物体内一种重要的矿质营养元素参与滇北球花报春对高温胁迫的反应。相关的生理指标和热害指数测定结果表明:Ca²⁺处理（20 mmol/L）能有效抑制高温胁迫引起的滇北球花报春幼苗叶片内丙二醛含量的快速积累,促进叶片内游离脯氨酸含量的积累,同时提高和保护Ca²⁺-ATPase酶的活性,并使热害指数降低近40%。Ca²⁺处理这种化学措施能从一定程度上提高滇北球花报春的耐热性。5.采用40种引物组合筛选到滇北球花报春对照和高温胁迫间差异明显的条带111条,回收到单一性条带70条,其中有5个差异片段经过Northern杂交验证。5个差异片段在GenBank中进行序列同源性比对后发现有2个差异片段与土豆、拟南芥和烟草等植物的热激蛋白具有较高的同源性,但其结构和功能还需要进一步研究确定。更多还原

【Abstract】 Primula is a famous alpine flower distributing throughout the moister and cooler regions of the northern hemisphere. By far the largest concentration of species is found in the great chain of Himalaya and western China, especially Hengduan Mountains in southwest China is the modern distribution and diversification centre of the genus Primula, and of which Yunnan Province has more than 120 species. Most species are beautiful and many species have proved to be great garden or ornamental flowers. The flower trade of Primula creates hundreds of millions dollars annually in American. While the study in China is far behind, and the Flora Reipublicae Popularis Sinicae on Primulaceae was completed until the early 1990s. In the past twenty years, the germplasm resources of Primula were destructed due to environmental factors and human factors （infrastructure construction, tourism development and so on）. And as the genus Primula is mainly located in higher altitude mountain regions, the characteristic of low heat resistence restricts its introduction and horticultural development process in China. Based on related documents and specimens, a field surney was carried out in northwest Yunnan Province. One representative Primula species, P. denticulata ssp. sinodenticulata was studied for the analysis of genetic diversity and establishment of conservation strategy.The main conclusions are as follows:1.During 3 years field survey in northwest Yunnan Province, 55 Primula species （including sub-species and varieties） belonging to 18 groups were located, which account for 50% of the species in Yunnan Province. The characteristics of Primula germplasm resources in this region were as follows: （1） species diversity was abundant and population density was variant, （2） distributed widely, habitats were complicated and various, （3） vertical distribution pattern appeared definite regularity. 2. According to comprehensive quantitative evaluation on ornemantal traits and ecological factors, 15 Primula species including P. sinolisteri, P. bathangensis, P. polyneara, P. forrestii, P. poissonii, P. wilsonii, P. chungensis, P. sikkimensis, P. faberi, P.sinopurprea, P. monticola, P. denticulate ssp.sinodenticulata, P. vialii, P. pinnatifida, P. deflexa have higher introduction value.3. 10 natural populatons of Primula denticulata ssp.sinodenticulata were located in Yunnan Province and 6 populatons in the region of Kunming, Song Ming, Yi Mun, Yuxi, Lijiang and Lushui were dismissed. And compared with to the records, population number and distribution area both had a certain degree of reduction, and its natural distribution had been fragmented. 300 copies pf these plants were chosed for initial core collection. The sampling strategy of core collection was established based on genetic diversity of morphological and AFLP markers, which includes （1） conserving 5 populations, （2） genotype conservation way by sampling 27～31 genotypes for each population.4.Ca2 + participated in the response of Primula denticulata ssp.sinodenticulata to heat stress as an important mineral nutrients. Results of related physiological indicators and heat harm index showed: Ca2 + treatment （20 mmol / L） could effectively inhibit the MDA’s rapid accumulation in leaves of this plant and improve increase of proline. Besides the treatment could also improve and maintain the activity of Ca²⁺-ATPase enzyme, and decrease the high harm index in 40 %. Chemical treatment of Ca2 + could improve the resistance of Primula denticulata ssp. sinodenticulata to heat stress in some extent.5. The technology of mRNA differential display was employed to isolated heat stress responsive genes in the leaves of Primula denticulata ssp.sinodenticulata. More than 1000 cDNA segments were amplified and 111were differeantial display. There were 5 cDNA segments only expressed in the state of heat stress. In the isolated segments, HSR4 shared 62% homology with HSP68 protein from Solanum tuberosun, 55% homology with mtHSC70-2 protein from Arabidopsis thaliana. HSR5 shared 70% homology with HSP90 protein from Nicotiana tabacum. Their expressions were possible involvement in the heat stress response, but their structure and function need to be further studied.更多还原