Assessing population genetic structure and variability with RAPD data: Application to Vaccinium macrocarpon (American Cranberry)

A method for estimating and comparing population genetic variation using random amplified polymorphic DNA (RAPD) profiling is presented. An analysis of molecular variance (AMOVA) is extended to accomodate phenotypic molecular data in diploid populations in Hardy-Weinberg equilibrium or with an assumed degree of selfing. We present a two step strategy: 1) Estimate RAPD site frequencies without preliminary assumptions on the unknown population structure, then perform significance testing for population substructuring. 2) If population structure is evident from the first step, use this data to calculate better estimates for RAPD site frequencies and sub-population variance components. A nonparametric test for the homogeneity of molecular variance (HOMOVA) is also presented. This test was designed to statistically test for differences in intrapopulational molecular variances (heteroscedasticity among populations). These theoretical developments are applied to a RAPD data set in Vaccinium macrocarpon (American cranberry) using small sample sizes, where a gradient of molecular diversity is found between central and marginal populations. The AMOVA and HOMOVA methods provide flexible population analysis tools when using data from RAPD or other DNA methods that provide many polymorphic markers with or without direct allelic data.     

A genetic linkage map of Theobroma cacao L.

A linkage map of the cocoa genome comprising 193 loci has been constructed. These loci consist of 5 isozymes, 101 cDNA/RFLPs, 4 loci from genes of known function, 55 genomic DNA/RFLPs and 28 RAPDs. A population of 100 individuals derived from a cross between two heterozygous genotypes was used. Segregation analyses were performed with the JoinMap program. Ten linkage groups, which putatively correspond to the ten gametic chromosomes of cocoa, were identified. The map covers a total length of 759 cM with a 3.9 cM average distance between 2 markers. A small fraction (9%) of the markers deviated significantly from the expected Mendelian ratios.     

不同地区根霉分离株的RAPD多样性分析

为了研究微生物的分子多样性,在获得系列米根霉分离物的基础上,应用经筛选的13条随机引物,对来自不同地区的18株米根霉分离物进行了RAPD分析,聚类结果表明,18株分离物在0.75的相异水平上可以聚为6个群体,其分子多样性同地理来源之间具有一定的相关性。     

濒危植物浮叶慈姑遗传多样性的RAPD分析

采用RAPD分子标记技术,对内蒙古二卡河(EK)和黑龙江库都尔(KDE)的浮叶慈姑居群进行了遗传多样性分析。 从60个随机引物中筛选出14个引物对2个居群共48个样品进行扩增,得到93条清晰的条带,其中70条具有多态性,即物 种水平上的多态位点百分率(PPB)为75.27%。POPGENE分析结果表明:二卡河居群的PPB为66.67%,库都尔居群的PPB 为72.04%。两个居群间基因分化系数(Gst)为0.0628,即遗传变异有很大一部分是来自居群内(93.72%)。结合相关的分析 表明生态学因素可能是浮叶慈姑濒危的主要原因。     

中国狗牙根(Cynodon dactylon)优良选系的RAPD分析

对采自中国不同地区的6份狗牙根[Cynodon dactylon (L.) Pers.]优良选系进行了RAPD分子标记实验.选用15个引物共扩增出438条带,平均每个引物扩增出29条带,其中多态性带415条,多态性位点百分率达到59.02%～75.49%.各选系间遗传相似性系数差异较大(0.408～0.672),说明各选系间在DNA水平上存在着丰富的遗传多样性,与新品种C106(爬地青)相比,各选系还存在一定的改良空间.实验结果也表明,RAPD分子标记可成功地用于中国狗牙根优良选系遗传多样性的研究及品种鉴定.     

宽叶泽苔草居群内遗传多样性研究

(武汉大学生命科学学院植物系统学与进化生物学研究室,武汉430072)摘要:采用RAPD分子标记对宽叶泽苔草(Caldesiagrandis)湖南浪畔湖居群30个家系共180个子代样品进行了居群内家系间以及家系内的遗传多样性分析。从100个随机引物中筛选出12个有效引物,共产生112条带,其中79条表现出多态性,占总条带数的70.5%。宽叶泽苔草各个家系多态位点百分率(PPB)在7.1%–40.2%之间。各个家系基因多样性范围在0.02–0.14之间(家系水平的基因多样性为0.10),Shannon指数的范围在0.04–0.21之间(家系水平的Shannon指数为0.18)。对宽叶泽苔草30个家系的AMOVA分析结果显示,家系间的基因分化系数Gst=0.231,即总的变异中有23.1%的变异存在于家系间,家系内的遗传变异占总遗传变异的76.9%,家系间和家系内变异均极显著(P<0.001)。采用UPGMA法对宽叶泽苔草30个家系180个子代进行聚类的结果表明:同一家系的子代个体并不能完全聚在一起。基于家系间的遗传分化系数Gst对宽叶泽苔草30个家系间的基因流进行估测,结果显示:Nm=0.83,表明宽叶泽苔草各个家系之间有较高的基因流。     

杜鹃属基因组DNA提取及RAPD的检定

以杜鹃花属常绿杜鹃亚属井冈山杜鹃树叶为材料,分别采用修改的CTAB法、碱裂解法、SDS法提取基因组DNA。实验表明,三种方法所提的DNA纯度及产率有差别,从综合结果看,以CTAB法最好,其所提到的DNA大小与λDNA（21kb）相似,经检验该法适合杜鹃花属植物总基因组DNA的提取,所得DNA不需纯化就可直接用于RAPD分析。     

卡瓦胡椒及胡椒的RAPD聚类分析

目的：通过RAPD分析搞清楚卡瓦胡椒与胡椒及胡椒属其它近缘野生种的亲缘关系。方法和结果：采用随机引物80条进行筛选,用从80条随机引物中入选的20条引物,对卡瓦胡椒和胡椒属共计28份材料进行RAPD扩增,均能产生清晰的扩增谱带。重复1—2次,结果稳定可靠。20个引物共扩增出170个条带,其中多态性条带有20条,占总扩增条带数的12％。RAPD分析结果显示在相似系数0．36处对28份种质可划分为6个类型,其中卡瓦胡椒被单独聚为一类,说明卡瓦胡椒与胡椒及其它近缘野生种的亲缘关系有一定的距离。     

顽拗植物龙眼基因组DNA提取方法的研究

为从顽拗植物龙眼(Dimocarpus longan Lour.)叶片中获得可供后续分子生物学操作的基因组DNA．针对其组织细胞内富含多酚、多糖、单宁及色素等物质的特点,采用改进的CTAB法和SDS法,即在核裂解之前先破碎细胞．将存在于细胞质中的次生物质去除后再裂解细胞桉．结合其它一些改进措施．提取到的DNA沉淀呈纯白色．极易溶解于TE中。两种改进方法的OD260和OD280比值分别达到1.82和1．73,其鲜叶基因组DNA产量分别为103ug/g和127ug／g：RAPD扩增条带清晰,丰富．完全满足后续分子生物学操作的要求,其中改良CTAB方法效果更为理想,与之埘比．传统的CTAB法和SDS法提取到的DNA沉淀呈浅黄色甚至红褐色,很难被TE溶解,其OD260和OD280比值均低于1．5,也得不到扩增产物。