烟草RAPD双引物与单引物PCR扩增多态性效率的比较分析

以两个亲缘关系较远的烟草（Nicotiana tabacum）品种台烟7号和白肋21为材料（GS=0.58）,研究了单引物扩增和双引物聚合扩增对RAPD-PCR分子标记多态性的影响。结果显示：双引物反应能够比单引物反应扩增出更多的多态性片段,双引物在白肋21和台烟7号中扩增出的多态性片段总数是单引物反应扩增出的多态性片段总数的6.6倍。因此,双引物RAPD的运用有助于提高烟草多态性分子标记的有效性。     

利用RAPD法对虹鳟鱼基因组DNA多态性研究初报

本文通过利用RAPD（随机扩增多态DNA）方法,选用20种10bp的随机引物（OPC－01－OPC—20）,用于虹鳟鱼基因组DNA多态性分析,其中发现一种引物（OPC－02）在虹鳟鱼群体中能扩增出多条带,说明该引物能反映其基因组DNA的多态性,可用于检测出不同品系间的差异。     

红麻及其近缘种的RAPD分析

利用随机扩增多态性DNA（RAPD）技术分析了木槿属（Hibiscus)Furcaria组中纤维作物红麻（H.cannabinus)及其6个近缘种植物的25份材料,用筛选出的16个引物扩增出192个RAPD条带,它们表现出丰富的多态性。根据得到的RAPD指纹图谱,计算其Nei氏相似系数和遗传距离。并构建了它们的系统树。结果表明,25份材料可划分为7个组。H.penduriformis和H.calyphyllus两个种为一组;红麻种分为两个组,一组为栽培品种,另1组为野生型材料;其余4个种各成为1组。玫瑰麻（H.sabdariffa)和金线吊芙蓉（H.radiatus)的关系较近,而且两者与红麻的亲缘关系也较近,而其它四个种与红麻的关系较远。H.trionum与其它六个种的关系较远。     

BALB/c小鼠遗传质量检测的新方法

目的比较随即扩增多态性方法（RAPD）、微卫星方法（STR）与生化标记方法对近交系小鼠遗传质量检测的差异,为近交系动物遗传质量控制提供一种分子生物学方法。方法提取近交系小鼠BALB/c基因组DNA,用6条RAPD引物和20对STR引物对其进行PCR扩增,用生化标记法检测13个位点。结果在6条RAPD引物中,引物2（p2）、引物3（p3）、引物5（p5）和引物6（p6）这四条引物扩增的条带出现差异,表现为不同的RAPD图谱;在20对STR引物中,引物2、4、10和11,这四对引物扩增的条带出现差异,表现为不同的STR图谱;13个生化标记位点中,过氧化氢酶-2（Ce-2）等6个生化位点发现杂合基因。结论RAPD和STR可用于验证生化标记方法的实验结果,并用于保证近交系动物的遗传质量。     

利用SSR与RAPD分子标记评估甘蔗品种的遗传多样性

利用SSR与RAPD两种分子标记对美国、中国台湾以及中国大陆不同甘蔗育种单位选育的甘蔗品种或亲本材料的遗传多样性进行评估。其中19对SSR引物共扩增出87条带,多态性带为84条,多态性比例为96.55%,扩增出的条带数范围为2~8条,平均每对引物扩增出4.58条带,引物的PIC值范围为0.34~0.93,平均0.64。21条RAPD引物共扩增出184条带,扩增条带数范围为3~16,平均每条引物扩增8.76条带,其中多态性带为184,多态性比例为100%,引物PIC范围为0.53~0.97,平均0.86。结果表明,两种分子标记都能较好的评估甘蔗品种的遗传多样性。     

利用RAPD和ISSR分子标记分析怀地黄种质遗传多样性

用RAPD与ISSR技术对怀地黄的8个品种和2个脱毒品系进行了种质遗传多样性分析。分别从80条RAPD引物和44条ISSR引物中筛选出适合怀地黄种质分析的17条RAPD引物和10条ISSR引物,用于RAPD和ISSR分析。17条RAPD引物共扩增出177条带, 多态性位点数为109; 多态性位点比率为61.58%;平均多样性指数(I)为0.3135;每个位点的有效等位基因数（Ne）是1.3641; 10条ISSR引物共扩增出110条带. 多态性位点数为79; 多态性位点比率为71.58%;平均多样性指数(I)为0.3577;每个位点的有效等位基因数（Ne）是1.4037。 基于扩增条带数据库建立了各自的Jaccard遗传相关系数矩阵,构建了相似的分子树状图,将10个供试材料分为2类:一类群含组培85.5、大田85.5、组培9302、大田9302、金状元和金白6个材料;另一类群含北京1号、大红袍、地黄9104和野生地黄4个材料。两种分子标记的分析结果呈极显著正相关(r＝0.649)。结果表明,RAPD与ISSR标记适合于怀地黄种质遗传多样性分析,ISSR标记技术是一种多态性和重复性优于RAPD技术的实用技术。     

亚麻遗传多样性的RAPD分析

从600个随机引物筛选出28个扩增稳定性较好的引物,对18份来自不同国家和地区的亚麻资源遗传多态性进行RAPD分析。结果表明：共扩增出条带529,其中多态性条带201,总的多态性百分率（PPB）为38．0%。用NTSYSpc（2．10）软件进行UPGMA聚类分析,18个亚麻品种遗传距离为0．0469～0．1332之间,可分3大类,其中红木5号与匈牙利5号亲缘关系最近,ABYSSINIA（BROWN）和匈牙利5号遗传距离相差最显著,达到0．1332。     

满天星试管苗与其玻璃化苗的RAPD指纹图谱分析

采用分离群体分组分析法(BSA),用100个随机引物对满天星的正常苗和玻璃化苗进行RAPD分析的结果表明,7个随机引物扩增出多态性差异条带。再用上述7个引物分别对试管苗及其玻璃化苗个体进行DNA的PCR扩增的结果显示,引物J20在2种苗中出现差异条带。     

云南普通野生稻遗传多样性和亲缘关系

野生稻（Oryza rufipogon）是稻属的重要组成部分,具有许多优良性状,是水稻遗传改良的天然基因库。本研究通过对形态学性状的观测,及ISSR和RAPDUPGMA聚类分析,将云南普通野生稻划分为4个类型,即元江类型、景洪紫杆直立型、景洪绿杆直立型和景洪匍匐型。在供试材料中筛选到具有多态性的ISSR和RAPD引物各11个,ISSR引物扩增出多态带113条,多态性条带比率（PPB）为82．26％,RAPD引物共扩增出多态性条带76条,PPB值为76．77％,两种分子标记的分析结果呈极显著正相关（r=0.951）。此外UPGMA聚类结果表明,云南普通野生稻不同类型与其它地区普通野生稻之间的遗传亲缘关系差异明显。     

应用RAPD和ISSR标记对24份花生栽培种材料进行遗传多样性分析

利用RAPD引物和ISSR引物分析我国24份花生栽培种材料的遗传多样性.结果表明:所选的RAPD引物和ISSR引物中分别有13条引物和10条引物扩增出了清晰并可重复的条带,共扩增出123条带和87条带,平均每条引物扩增出9.5条带和8.7条带,其中多态性带分别占条带总数的47.15％和57.47％,平均每条引物扩增出4...     

Identification and genetic mapping of random amplified polymorphic DNA (RAPD) markers to the sheep genome

The random amplified polymorphic DNA (RAPD) assay utilizes the polymerase chain reaction (PCR) and short primers of arbitrary nucleotide sequence to amplify DNA. In this study, the RAPD assay was used to identify and map polymorphic markers in the AgResearch International Mapping Flock (IMF) sheep pedigrees. Sires and dams of eight of the full-sib IMF pedigrees were screened with 131 different 10-mer oligonucleotide primers. An average of 85 RAPD polymorphisms was identified between each parental pair, and 53 markers were contributed to the AgResearch IMF collaboration. Forty-five of the RAPD markers were mapped in the AgResearch IMF genetic linkage map, and at least one marker was located on 17 of the 26 autosomes and both sex chromosomes. Three lines of evidence were used to check for the homology of scored polymorphisms in different pedigrees, pedigree evaluation, segregation analysis, and Southern blot analysis. These results demonstrate that the RAPD assay is a powerful approach for identifying polymorphisms that can be used as markers for constructing a sheep genetic linkage map. Received: 5 October 1995 / Accepted: 16 April 1996     

Comparative studies of genetic diversity in kenaf (Hibiscus cannabinus L.) varieties based on analysis of agronomic and RAPD data

Kenaf (Hibiscus cannabinus L.) is a fiber crop classified in the genus Hibiscus (Malvaceae), and has a great potential for its multipurpose utilization, in addition to its traditional usage. Varietal identification of kenaf is always problematic and knowledge on genetic diversity of kenaf varieties is also limited, which significantly hindered our effective utilization and conservation of the valuable kenaf germplasm. In order to find a proper method for identifying kenaf varieties and studying their variation, morpho-agronomic characters and random amplified polymorphic DNA (RAPD) markers were analyzed among 14 kenaf varieties commonly used in Japan. Data from morphological analysis showed that the included kenaf varieties could be divided into three major groups. The characters, such as middle stem diameter, whole stalk weight, and days to 50% flowering, are highly responsible for the variation of the kenaf varieties, but it is difficult to identify individual varieties merely by the morpho-agronomic characters. On the other hand, clearly separation of the kenaf varieties was achieved based on the RAPD variation patterns. Genetic relationship of the kenaf varieties can also be traced through the analysis of RAPD and morph-agronomic variation. It is concluded from the present study that RAPD analysis is an effective tool in identifying of kenaf varieties and determining their genetic relationships, particularly when combined with the analysis of morpho-agronomic characters.     

Genetic Linkage Maps of Eucalyptus Grandis and Eucalyptus Urophylla Using a Pseudo-Testcross: Mapping Strategy and Rapd Markers

We have used a ``two-way pseudo-testcross' mapping strategy in combination with the random amplified polymorhic DNA (RAPD) assay to construct two moderate density genetic linkage maps for species of Eucalyptus. In the cross between two heterozygous individuals many single-dose RAPD markers will be heterozygous in one parent, null in the other and therefore segregate 1:1 in their F(1) progeny following a testcross configuration. Meiosis and gametic segregation in each individual can be directly and efficiently analyzed using RAPD markers. We screened 305 primers of arbitrary sequence, and selected 151 to amplify a total of 558 markers. These markers were grouped at LOD 5.0, θ = 0.25, resulting in the maternal Eucalyptus grandis map having a total of 240 markers into 14 linkage groups (1552 cM) and the paternal Eucalyptus urophylla map with 251 markers in 11 linkage groups (1101 cM) (n = 11 in Eucalyptus). Framework maps ordered with a likelihood support >/=1000:1 were assembled covering 95% of the estimated genome size in both individuals. Characterization of genome complexity of a sample of 48 mapped random amplified polymorphic DNA (RAPD) markers indicate that 53% amplify from low copy regions. These are the first reported high coverage linkage maps for any species of Eucalyptus and among the first for any hardwood tree species. We propose the combined use of RAPD markers and the pseudo-testcross configuration as a general strategy for the construction of single individual genetic linkage maps in outbred forest trees as well as in any highly heterozygous sexually reproducing living organism. A survey of the occurrence of RAPD markers in different individuals suggests that the pseudo-testcross/RAPD mapping strategy should also be efficient at the intraspecific level and increasingly so with crosses of genetically divergent individuals. The ability to quickly construct single-tree genetic linkage maps in any forest species opens the way for a shift from the paradigm of a species index map to the heterodox proposal of constructing several maps for individual trees of a population, therefore mitigating the problem of linkage equilibrium between marker and trait loci for the application of marker assisted strategies in tree breeding.     

RAPD和SSR两种标记构建的中国对虾遗传连锁图谱

利用RAPD和SSR分子标记结合拟测交策略,对中国对虾(Fenneropenaeuschinensis)“黄海1号”雌虾与野生雄虾作为亲本进行单对杂交产生的F1代,采用RAPD和SSR两种分子标记技术初步构建了中国对虾雌、雄遗传连锁图谱。对460个RAPD引物和44对SSR引物进行筛选,共选出61个RAPD引物和20对SSR引物,用于对父母本和82个F1个体进行遗传分析。共得到母本分离标记146个(RAPD标记128个,微卫星标记18个)和父本分离标记127个(RAPD标记109个,微卫星标记18个)。雌性图谱包括8个连锁群、9个三联体和14个连锁对,标记间平均间隔为11·28cM,图谱共覆盖1173cM,覆盖率为59·36%;雄性图谱包括10个连锁群、12个三联体和7个连锁对,标记间平均间隔为12·05cM,图谱共覆盖1144·6cM,覆盖率为62·01%。中国对虾遗传图谱的构建为其分子标记辅助育种、比较基因组作图及数量性状位点的定位与克隆奠定了基础。     

珍稀濒危植物天目木兰（Magnolia amoena）遗传多样性的RAPD分析

运用随机扩增多态DNA(RAPD)技术,对天目木兰(Magnolia amoena)居群的遗传多样性进行了研究．从40个10-mer随机引物中筛选出14个能得到清晰、稳定扩增带的引物进行扩增,14个引物共检测了94个位点,其中多态性位点为23,占24．4％,计算了12个居群之间的遗传相似度和遗传距离,并运用UPGMA法进行了聚类分析,结果显示相同严地个体间(居群内)的遗传距离较小,遗传多样性水平很低;不同产地个体间(居群间)遗传距离较大,遗传多样性水平较前者高,即天目木兰个体间遗传多样性水平与它的地理分布有关,天目木兰总体较低的遗传多样性是导致它濒危的原因之一。     

Construction of Molecular Linkage Map in Masson Pine Using RAPD Markers and Megaga metophytes from a Single Tree

A set of 300 random ohgonucleotide primers was screened for random amplified polymorphic DNA (RAPD) fragments within a sample of 79 megagametophyte DNAs of a single masson pine ( Pinus massoniana Lamb. ) to generate RAPD markers and construct the molecular linkage map for masson pine. 64 repeatable RAPD markers segregated in a 1 present to 1 absent ratio. Through multipoint linkage analysis, 47 markers were assigned to 13 different linkage groups(pairs). It covered a tdtal genetic distance of over 692.5 cM (centimorgan). The average distance between markers was 14.7 cM. It supplied a linkage framework for a more saturated linkage map of masson pine.     

Evaluation of inter-simple sequence repeat analysis for mapping in Citrus and extension of the genetic linkage map

Inter-simple sequence repeat (ISSR) analysis was evaluated for its usefulness in generating markers to extend the genetic linkage map of Citrus using a backcross population previously mapped with restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD) and isozyme markers. ISSR markers were obtained through the simple technique of PCR followed by analysis on agarose gels, using simple sequence repeat (SSR) primers. Optimization of reaction conditions was achieved for 50% of the SSR primers screened, and the primers amplified reproducible polymorphic bands in the parents and progeny of the backcross population. Mendelian segregation of the polymorphic bands was demonstrated, with an insignificant number of skewed loci. Most of the SSR primers produced dominant loci; however co-dominance was observed with loci derived from three primers. A new genetic map was produced by combining the segregation data for the ISSR markers and data for the RFLP, RAPD and isozyme markers from the previous map and creating genetic linkages among all the markers using JoinMap 2.0 mapping software. The new map has an improved distribution of markers along the linkage groups with fewer gaps, and marker order showed partial or complete conservation in the linkage groups. The incorporation of ISSR markers into the genetic linkage map demonstrates that ISSR markers are suitable for genetic mapping in Citrus. Received: 3 February 2000 / Accepted: 12 May 2000     

利用RAPD标记和单株树大配子体构建马尾松的分子标记连锁图谱

利用300个随机的寡核苷酸引物和马尾松(Pinus massoniana Lamb.)种子的胚乳(大配子体)产生的随机扩增多态性DNA(RAPD)分子标记,进行马尾松标记连锁图谱的构建。经筛选共获得64个稳定的RAPD标记,利用多点连锁分析,其中的48个标记分属于13个不同的连锁群(连锁对),该图谱覆盖的基因组总长度约为692.5cM(centimorgan),标记间平均距离约为14.7cM,它为马尾松连锁图谱的构建提供了一个连锁框架。     

Genetic variation in monoploids of diploid potatoes and detection of clone-specific random amplified polymorphic DNA markers

Randomly amplified polymorphic DNA (RAPD) markers have been used to study the genetic variation among androgenetic monoploids of diploid Solanum species. Cluster analysis of pairwise genetic distances was used to construct a genetic relationship among anther donor and anther-derived potato plants. The clustering based on Rogers' distances resembled classifications based on parental origins and hybrid combinations. Six of the 32 RAPD primers used resulted in the selective amplification of DNA fragments which were polymorphic between the two S. phureja parental clones, 1.22 and A95. It should be possible to construct a genetic linkage map, without making crosses, using monoploids derived from a single heterozygous diploid clone and RAPD markers.