湖北省油茶优良单株遗传多样性及品质初步分析

Analysis on Genetic Diversity and Quality of Camellia Oleifera Resources in Hubei Province

作者: 专业:林木遗传育种 导师:涂炳坤 年度:2010 学位:硕士  院校: 华中农业大学

Keywords

Hubei province, Genetic diversity, Camellia oleifera Economic characters, Fatty acid composition, ISSR, Gas chromatography

        油茶(Camellia oleifera Abel.)是指山茶科(Theaceae)山茶属(CamelliaL.)油脂含量较高且有栽培经济价值的一类植物的总称。本研究从湖北省的东南西北四个区域的不同地区采集了无病虫害且生长旺盛的优良单株新鲜叶片,进行ISSR遗传多样性分析。同时针对25个表现良好的单株果实进行脂肪酸分析,为下一步选优工作提供理论依据。主要结果如下:(1)从99条ISSR引物中筛选出16条多态性带数且谱带亮度等方面均表现良好的引物,用这16条引物对110个油茶叶样的基因组DNA进行PCR扩增,共得到221条扩增谱带,其中多态性条带218条,多态性百分率(PPB)达到99.09%。(2)分别采用多态位点百分率(PPB)、有效等位基因指数(AE)、观测等位基因指数(A0)、Ne’基因多样性指数(HE)以及Shannon’s多态信息指数来衡量油茶的遗传多样性,结果表明,湖北省不同产地间的油茶的遗传多样性水平较高:湖北省油茶的物种水平遗传多样性达到99.09%,群体多态位点百分比达到了61.49%,Nei’s遗传多样性指数为0.2776,Shannon’s多态信息指数为0.4291。(3)七个主要县市的遗传多样性比较:松滋地区的各项指标均为最高,五峰地区的多态性百分率和观测等位基因(AO)指数为最低,武汉狮子山的有效等位基因指数(AE)、Nei’s基因多样性指数和Shannon’s信息指数均为最低。松滋地区的多态性百分率为85.45%,观测等位基因指数(AO)为1.8545,有效等位基因指数(AE)为1.3852,Nei’s基因多样性指数为0.2423,Shannon’s信息指数为0.3795。五峰地区的多态性百分率为52.73%,观测等位基因(AO)指数为1.5273。武汉狮子山的有效等位基因指数(AE)为1.2539、Nei’s基因多样性为0.157和Shannon’s信息指数为0.2438。(4)根据群体间的Nei’s遗传一致度(I)和Nei’s遗传距离(D),应用NTSYS2.1软件,采用UPGMA法构建聚类图,结果发现:麻城、阳新、新洲、五峰、谷城和武汉六个地区的个体基本上可以聚集在一起,而松滋地区的不同个体不能按地区聚集在一起间,遗传距离相差很大,这可能是因为其资源是由不同地方引进的原因导致的。(5)油茶的主要经济性状与鲜籽含油率相关性大小为:干仁含油率>鲜籽出仁率>种仁含水率>冠幅单面积产量>单株产量>冠幅。其中与之呈显著正相关的是鲜籽出仁率和干仁含油率,相关系数分别为0.622、0.928。其他指标虽然与之呈负相关,但不显著。(6)油茶的鲜籽含油率为14.84%~36.41%,表明地区间有较大的差异。(7)油茶脂肪酸的成分主要是油酸、亚油酸、棕榈酸和硬脂酸,不同采样地点的单株脂肪酸组成有一定的差异。其中不饱和脂肪酸总量范围为76.30%~91.04%,平均含量为88.86%,变动系数为3.51%;油酸含量范围为65.70%~84.48%,平均含量为79.85%,变动系数为5.14%;亚油酸含量范围为6.14%~13.37%,平均含量为9.63%,变动系数为23.01%;棕榈酸含量范围为6.61%~11.01%,平均含量为7.97%,变动系数为11.88%。(8)油茶种子各脂肪酸组成成分的含量之间存在显著相关性,其中油酸与亚油酸含量存在极显著负相关,相关系数-0.631,而二者含量之和相对稳定。
    Oil-Tea belongs to Camellial.L.of Theaceae which is a unique and important edible woody oil plant in southern China and which has high economic value and high oil content. It is one of the four famous oil plants in the world. In this study, the excellent healthy fresh Plant leaves of vigorous growth were collected from different areas of Hubei Province and the molecular genetic diversity was analyzed. Meanwhile, the main results are as follows:(1) Sixteen Polymorphic Primers with good, clear bands were got, screening from seventy-five ISSR primers. These sixteen pairs of primers were used to amplification respectively the genomic DNA of 110 different Camellia oleifera individuals and a total of 220 bands were amplified with 218 polymorphic bands and percentage of polymorphic bands (PPB) 99.09%.(2) Genetic variation of Camellia oleifera was measured respectively by the Percentage of Polymorphic bands(PPB), the number of alleles observed(Ao), effective gene(AE), Nei’s genetic diversity(HE) and Shannon’s polymorphic information index. The results showed that Camellia oleifera populations had a high level of genetic diversity:The level of genetic diversity of species was relatively high, groups polymorphic loci percentage reached 99.09%, Nei’s degree of genetic diversity was 0.2776 and Shannon’s polymorphic Information index was 0.4291.(3) The genetic diversity of sample in 7 counties showed that Songzi exhibited the highest on the four indexes. The percentage polymorphic loci, observation allele(AO), effective allele(AE), Nei’ s genetic diversity and Shannon’s information index in SongZi were 85.45%,1.8545,1.3852, 0.2423 and 0.3795. In WuFeng the percentage polymorphic loci, observation allele(AO)were the lowest index. The values of the effective alleles, Nei’s genetic diversity and Information highest index were the lowest in Wuhan.(4) According to the Nei group’s genetic identity. UPGMA clustering of individual plans was constructed. Individuals from the same place gathered together basically and the different individuals in SongZi varied greatly. That may be considered that Camellia oleifera in SongZi were introduced from various places.(5) The order of the correlation between seed oil content and the main economic characterisitics is the seed oil content>the kernel content of seed>water content of kernel> output of unit crown area>yield per plant>crown width. Of them, there are significantly positive correlation between the kernel content of seed, oil content of dry kernel and the seed oil content, with the correlation coefficient being 0.622 and 0.928 respectively, while the correlation between the seed oil content and other characters is negative.(6) Oil content of fresh seed ranges from 14.84% to 36.41%, and there are great differences between the strains;(7) Fatty acid of Camellia oleifera oil is mainly composed of oleic, linoleum and palmitic acid, and there are still some differences among strains. The content of unsaturated acids ranges from 76.30% to 91.04%, the average being 88.86%, and the variation coefficient is 3.51%; the content of oleic acid ranges from65.70% to 84.48%, the average being 79.85%, and the variation coefficient is 5.14%; the content of linoleic acid ranges from 6.14% to 13.37%, the average being 9.63%, and the variation coefficient is 23.01%; the content of palmitic acid ranges from 6.61% to 11.01%, the average being 7.97%, and the variation coefficient is 11.88%.(8) There are significant correlations among the contents of each component of fatty acid; an extremely negative correlation is observed between the contents of oleic and linoleic acid, and the correlation coefficient is-0.631; however, the sum of the contents of oleic and linoleic acid is relatively stable.
        

湖北省油茶优良单株遗传多样性及品质初步分析

摘要7-9
Abstract9-10
缩略语表11-12
1 文献综述12-24
    1.1 油茶种质资源12-13
    1.2 生物学特性及分布13-14
        1.2.1 形态特征13
        1.2.2 生物学特性13
        1.2.3 地理分布13-14
    1.3 茶油的利用14-16
    1.4 湖北省油茶产业发展现状16
        1.4.1 油茶产业发展现状16
    1.5 分子标记研究背景16-21
        1.5.1 分子标记的概念界定17-18
        1.5.2 DNA分子标记的种类18-21
    1.6 油茶的研究进展21-23
        1.6.1 表形性状研究进展21
        1.6.2 油茶生物技术研究进展21-23
        1.6.3 油脂成分分析研究进展23
    1.7 研究的目的与意义23-24
2 材料与方法24-30
    2.1 材料24-25
    2.2 主要试剂、仪器及其来源25-26
    2.3 试验方法26-30
        2.3.1 基因组总DNA的提取与质量检测26-27
        2.3.2 引物筛选27-28
        2.3.3 ISSR反应条件的优化和反应体系的建立28
        2.3.4 ISSR扩增产物检测28
        2.3.5 谱带统计及数据分析28-30
3 结果分析30-39
    3.1 ISSR扩增产物的多态性分析30-33
    3.2 16条引物扩增结果33
    3.3 7 个县市的油茶遗传多样性33-35
    3.4 110份油茶DNA基于UPGMA法的聚类分析35-37
    3.5 ISSR标记的主坐标分析37-39
4 讨论39-41
    4.1 取样对遗传变异的影响39
    4.2 湖北省油茶不同个体间的遗传关系39-40
    4.3 湖北省主要县市群体间的遗传关系40-41
第二部分 25个复选油茶单株含油率和脂肪酸组成的初步研究41
1 材料与方法41-43
    1.1 材料41
    1.2 试验地概况41
    1.3 试验试剂及仪器41-42
    1.4 试验方法42-43
        1.4.1 油茶主要性状指标测定方法42
        1.4.2 测定样品制取42
        1.4.3 脂肪酸组成分析42-43
    1.5 统计分析方法43
2 结果与分析43-50
    2.1 各性状间的相关关系43-45
    2.2 含油率分析45-46
        2.2.1 不同单株的种子含油率45-46
        2.2.2 两不同产区种子含油率46
    2.3 油茶种子脂肪酸组分及含量比较分析46-49
    2.4 脂肪酸组成及其含量的初步定量分析49-50
        2.4.1 油酸49
        2.4.2 亚油酸49
        2.4.3 棕榈酸49
        2.4.4 硬脂酸49-50
        2.4.5 脂肪酸组成间的相关性50
3 结论与讨论50-53
    3.1 结论50-51
    3.2 讨论51-52
        3.2.1 关于油茶含油率的问题51
        3.2.2 关于脂肪酸组成51-52
        3.2.3 关于湖北省油茶产业发展52
    3.3 试验中的不足52-53
参考文献53-59
附录59-60
致谢60
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