Analysis of genetic diversity for wild and captive green peafowl populations by random amplified polymorphic DNA technique

The genetic diversity of the populations for 14 wild green peafowls (Pavo muticus) and 18 captive green pea-fowls was investigated by using the technology of random amplified polymorphic DNA (RAPD). Totally 161 and 166 ampli-fied bands were obtained by using 23 arbitrary primers to amplify the genomic DNA of wild and captive green peafowls re-spectively. The results showed that the average relative genetic distance of the wild and captive green peafowls popula-tions was 0.0555 and 0.1355, respectively, and difference of the average relative genetic distances between the two popu-lations was 0.1635. The Shannon diversity index for the wild and captive green peafowl populations was 0.4348 and 1.0163, respectively, which means that there exists significant difference in genetic diversity between the two populations, and the genetic diversity of wild green peafowl was low. The two populations originated from two different families according to analysis by the UPGMA method. This research can provide the theoretical basis for supervising genealogies management of peafowl populations.

Use of random amplified polymorphic DNA （RAPD） analysis to detect jujube witches＇ broom phytoplasmas

Jujube witches＇ broom is a devastating disease of Ziziphusjujube that occurs in various jujube regions of China. Nucleic acid extracted from midribs of samples collected from three jujube varieties （＂Suanzao＂, ＂Lajiaozao＂ and ＂Langzao＂） from symptomatic and asymptomatic shoots were tested by random amplified polymorphic DNA analyses. Using 13 different 10 and 11-bp random primers the amplification of jujube DNA was achieved from all the samples; AMI4 primer provided amplification of specific DNA fragments of about 400 bp, only from samples collected from symptomatic plants. No genetic variations in these varieties were identified using the other 11 arbitrary primers; only with primer AL07 it was possible to differentiate ＂Langzao＂ from the other two varieties tested. All the experiments were repeated 2 times and the results were consistent. Compared with PCR analyses with phytoplasma-specific primers, RAPD techniques resulted to be an alternative rapid and sensitive method for detecting jujube phytoplasmas presence in different jujube varieties.

Authentication and Genetic Origin of Medicinal <i>Angelica acutiloba</i>Using Random Amplified Polymorphic DNA Analysis

Some Angelica species are used for medicinal purposes. In particular, the roots of Angelica acutiloba var. acutiloba and A. acutiloba var. sugiyamae, known as “Toki” and “Hokkai Toki”, respectively, are used as important medicinal materials in traditional Japanese medicine. However, since these varieties have recently outcrossed with each other, it is difficult to determine whether the Japanese Angelica Root material used as a crude drug is the “pure” variety. In this study, we developed an efficient method to authenticate A. acutiloba var. acutiloba and A. acutiloba var. sugiyamae from each other and from other Angelica species/varieties. The random amplified polymorphic DNA (RAPD) method efficiently discriminated each Angelica variety. A. acutiloba var. sugiyamae was identified via a characteristic fragment amplified by the decamer primer OPD-15. This fragment showed polymorphisms among Angelica species/varieties. The unique fragment derived from A. acutiloba var. sugiyamae was also found in one strain of A. acutiloba var. acutiloba, implying that this strain arose from outcrossing between A. acutiloba var. acutiloba and A. acutiloba var. sugiyamae. This RAPD marker technique will be useful for practical and accurate authentication among A. acutiloba var. acutiloba, A. acutiloba var. sugiyamae, and their adulterants.

RAPD analysis of alfalfa DNA mutation via N^+ implantation

Germination capacity of alfalfa seeds under low energy N+ implantation manifests oscillations goingdown with dose strength. From analyzing alfalfa genome DNA under low energy N+ implantation by RAPD (RandomAmplified Polymorphous DNA), it is recommended that 30 polymorphic DNA fragments be amplified with 8 primersin total 100 primers, and fluorescence intensity of the identical DNA fragments amplified by RAPD is different be-tween CK and treatments. Number of different polymorphic DNA fragments between treatment and CK via N+ im-plantation manifests going up with dose strength.

Intra-specific relationships among Tibetan Eared-pheasants based on randomly amplified polymorphic DNA(RAPD)analysis

The Tibetan Eared-pheasant Crossoptilon harmani is a rare species native to China.A captive population has been established in the Beijing Zoo since 1999.In order to determine the kinship of the offsprings in 2001,randomly amplified polymorphic DNA(RAPD)was used to examine the parenthood of seven Tibetan Eared-pheasants in the Beijing Zoo.To amplify the genomic DNA of each individual,53 arbitrary primers were selected.The results of amplifications showed that 14 primers had clear and distinct RAPD patterns.Totally,226 amplified fragments were generated by RAPD in this study.Cluster analysis of the seven Tibetan Eared-pheasants indicated that all the four young birds had the same father(No.5 male).This study provides a practical method to determine the relationship of offsprings whose parents are unknown in birds.

Analysis of the Genetic Structure of Sclerotinia sclerotiorum （Lib.） de Bary Populations from Different Regions and Host Plants by Random Amplified Polymorphic DNA Markers

: The genetic diversity and genetic structure of a population of isolates of Sclerotinia sclerotiorum (Lib.) de Bary from different regions and host plants were investigated using the random amplified polymorphic DNA (RAPD) method with 20 random decamer primer pairs in order to provide some information on the phylogenetic taxa and breeding for resistance to sclerotinia stem rot. A minimum of three and a maximum of 15 unambiguously amplified bands were generated, furnishing a total of 170 bands ranging in size from 100 to 3 200 bp, corresponding to an average of 8.5 bands per primer pair. One hundred and four of these 170 bands (61.2%) were polymorphic, the percentage of polymorphic bands for each primer pair ranging from 0.0% to 86.7%. The genetic relationships among the isolates, based on the results of RAPD analysis, were examined. The genetic similarity of all selected isolates was quite high. At the species level, the genetic diversity estimated by Nei's gene diversity (h) was 0.197 and Shannon's index of diversity (I) was 0.300. The unweighted pair-group mean analysis (UPGMA) cluster analysis showed that most isolates from the same regions were grouped in the same cluster or a close cluster. The population of isolates from Hefei (Anhui Province, China) was more uniform and relatively distant to other populations. The Canadian population collected from carrot (Daucus carota var. sativa DC.) was relatively close to the Polish population collected from oilseed rape (Brassica napus L.) plants. There was no relationship between isolates from the same host plants. An analysis of molecular variance (AMOVA) revealed that the percentage of variance attributable to variation among and within populations was 50.62% and 49.38%, respectively. When accessions from China, Europe, and Canada were treated as three separate groups, the variance components among groups, among populations within groups, and within populations were ?0.96%, 51.48%, and 49.47%, respectively. The genetic differentiations among and within populations were highly significant (P < 0.001). Similarly, the coefficient of gene differentiation (Gst) in total populations calculated by population genetic analysis was 0.229 4, which indicated that the genetic variation among populations was 22.94%. The gene flow (Nm) was 1.68, which indicated that the gene permutation and interaction among populations was relatively high.

Genetic analysis of selected Sargassum fusiforme (Harvey) Setchell (Sargassaceae, Phaeophyta) strains with RAPD and ISSR markers

Since the 1980s,Sargassum fusiforme has been cultivated in Zhejiang,South China,and nowadays it becomes one of the important commercial seaweeds in China.With traditions of eating habits in the East Asian countries,this brown alga is used as food,because it contained functional oligo/polysaccharides and chemical components,and was regarded playing roles in antioxidant activities and regulating immunology.Through over 15 years’selection,breeding and cultivation,we obtained three strains with good traits and testified their characters during the production,which included the cultivars with high yield and other two good characters,either all the selected strains were applied in the Sargassum production.To avoid confusion during the selection and nursery,it was preferred to establish one fingerprint for distinguishing the Sargassum cultivars from different strains.Random amplified polymorphic DNA(RAPD)and inter-simple sequence repeat(ISSR)methods were adopted to analyze the genetic diversities of the selected S.fusiforme strains.With that,one fingerprint with RAPD markers was constructed,and one sequence characterized amplifi ed region(SCAR)marker to S.fusiforme was obtained.It is indicated that the applied fingerprint could be valid in S.fusiforme genetic and germplasm justification,and will be positive to molecular marker assistance in its selection and cultivation.

RAPD analysis of natural populations of Acanthopanax brachypus

Random Amplified Polymorphic DNA (RAPD) analysis is a new technology of molecular marking which has proved very powerful in detecting genetic diversity at the level of population. The genomic DNAs used in our experiment were extracted from fresh leaves taken from 59 individuals sampled from three natural populations in Yan An, Shanxi Province. Through more than 2,000 PCRs, deep-going RAPD analysis was carried out on DNA samples from 49 inviduals. The percentage of polymorphic RAPD loci found in these three populations were respectively 27.2 %, 18.6 % and 5.4 %; the average genetic distances within population, 0.055, 0.036 and 0.008; the average genetic distances between populations (Ⅰ-Ⅱ), (Ⅰ-Ⅲ) and (Ⅱ-Ⅲ), 0.105, 0.096 and 0.060. The genetic diversity of A. brachypus within and between populations was found, for the first time, to be rather poor,thus revealing innate factors as the cause contributing to its endangered status. In addition, our work also provides basic materials for elucidating the underlying cause of its endangerment and for its protection biology.

RAPD Analysis of Rosa laevigata Michx. from Different Origins

[ Objective] This study aimed to investigate the genetic diversity and phylogenetic relationship of 30 populations of Rosa laevigata Michx. from different origins. [ Method] Using RAPD molecular marker technique, eight effective primers were screened from 35 RAPD random primers for amplification. UPGMA clus- ter analysis was performed using NTSYSpc. ver. 2.02 software. [ Result] A total of 86 loci were amplified using the screened primers, including 84 polymorphic loci, indicating an average polymorphic percentage （PPB） of 97.67%. The similarity coefficients of 30 populations of R. laevigata Michx. ranged from O. 11 to O. 58, and these populations were clustered into groups A, B and C. [ Conclusion ] R. laevigata Michx. exhibits multiple genetic polymorphism. R. laevigata Michx. populations from Guizhou Province have closer genetic relationships.

RAPD-PCR Analysis on Genetic Relationships Between Cultivars of Tree Peony

Random amplified polymorphic DNA (RAPD) was used to analyze genetic polymophism of 35 Tree Peony cultivars with 7 different color groups. Thirty four primers amplified 418 DNA fragments and 337 polymorphic bands (80.6%), including specific DNA markers for 18 cultivars that could be used to differentiate cultivars. The UPCMA method was used to analyze the genetic relationship among cultivars. The results showed that 35 Peony cultivars could be divided into 2 cluster groups when using similarity criteria of 1.5, and into 4 cluster groups when using similarity criteria of 1.0. The result confirmed that the flower color has no relation to the genetic clusters and the Tree Peony cultivars originated from the same area has close genetic relationship. Therefore, genetic background has no large effect on the genetic relationship. The sequence based on polymorphic rate from high to low was Blue groups > Yellow groups > Bark red groups > Blake groups > White groups>Green groups>Red groups.

Micropropagation of loblolly pine by somatic organogenesis andRAPD analysis of regenerated plantlets

Organogenesis was induced in callus derived from mature zygotic embryos of six families (J-56, S-1003, E-22, E-311, E-440, and Mc) of loblolly pine (Pinus taeda L.) within 24 weeks of culture. Elongation of adventitious buds was achieved on TE medium supplemented with 0.5 mg·L?1 indole-3-butyric acid (IBA) and 1 mg/l 6-benzyladenine (BA). The most suitable medium for root formation proved to be TE medium supplemented with 0.5 mg·L?1 IBA, 2mg·L?1 BA and 0.5 mg/l gibberellic acid (GA3). 169 regenerated plantlets were transferred to a perlite: peatmoss: vermiculite (1∶1∶1) soil mixture, and 98 plantlets survived in the field. Total DNA was extracted from the needles of the regenerated plantlets of the six families of loblolly pine. Analysis of random amplified polymorphic DNA (RAPD) using 20 arbitrary oligonucleotide 10-mers, show that amplification products were monomorphic for all the plantlets of family J-56, S-1003, E-22, E-311, E-440, and Mc of loblolly pine. These results suggested that organogenesis can be used for clonal micropropagation of some families of loblolly pine.

Use of Random Amplified Polymorphic DNA Analysis for Economically Important Food Crops

The objective of this review is to summarize numerous studies on the use of the random amplified polymorphic DNA （RAPD） technique on rice, corn, wheat, sorghum, barley, rye, and oats to examine its feasibility and validity for assessment of genetic variation, population genetics, mapping, linkage and marker assisted selection, phylogenetic analysis, and the detection of somaclonal variation. Also we discuss the advantages and limitations of RAPD. Molecular markers have entered the scene of genetic improvement in different fields of agricultural research. The simplicity of the RAPD technique made it ideal for genetic mapping, plant and animal breeding programs, and DNA fingerprinting, with particular utility in the field of population genetics.

Genetic Differentiation Caused by Chromium Treatment in <i>Leersia hexandra</i>Swartz Revealed by RAPD Analysis

Randomly amplified polymorphic DNA (RAPD) technique was applied to assess the genetic variations and phylogenetic relationships in genetic differentiation within 4 Chromium-treatment Leersia hexandra. The fresh leaves of Leersia hexandra cultivated on the condition of chrome pollution and exogenous organic acids were used as experimental material. The genomic DNA of Leersia hexandra was extracted by using CTAB method. The results showed that different samples of Leersia hexandra exhibited DNA polymorphism when using the random primer S43, S51and S55 as the primers in the RAPD reaction. One specific DNA band about 1000 bp was found in the sample which treated with 10 mmol/L concentration EDTA when used the S43 primer to RAPD. The obvious differences between different EDTA-treatment levels suggest that EDTA has certain effects on enrichment to heavy metals of Leersia hexandra, it will be more favored to Leersia hexandra accumulation of chromium when EDTA concentration increased.

三系杂交稻亲本随机扩增多态性DNA(RAPD)分析

选用９个随机引物对３１份杂交水稻亲本材料进行了ＲＡＰＤ分析，共检测到６０条多态性带。聚类分析结果表明，所有供试材料可以被明确地区分。在９个随机引物中，有８个具有较高的多态性检测能力。以这８个引物为基础，选用任两个引物即可在任一对材料中检测出多态性的频率在９６１３％以上，而选用任３个引物则该频率在９９２１％以上。这显示了运用ＲＡＰＤ鉴定稻种具有简便、灵敏、高效的优点，在鉴定杂交稻种的实践中有着良好的应用前景。

栽培稻和普通野生稻基因组的随机扩增多态性DNA（RAPD）初步分析

用３５个随机引物对５个籼稻品种、５个粳稻品种和２７份中国普通野生稻进行了ＲＡＰＤ分析。６０％以上的引物能在籼稻和粳稻基因组间显示差异；中国普通野生稻与栽培稻的差异主要表现在与籼稻的不同，绝大部分供试野生稻的ＲＡＰＤ带型与粳稻的相同。这说明多数中国普通野生稻偏粳，但也存在偏籼的普通野生稻。

RAPD分析氮离子注入甜菊种子后的幼苗基因组DNA变异

应用ＲＡＰＤ 技术检测经低能氮离子注入甜菊纯系种子引起的幼苗基因组ＤＮＡ 变异。筛选出ＯＰＪ系列中的１５ 种引物对实验及对照基因组ＤＮＡ 进行了ＰＣＲ 扩增，共获扩增片段１０３ 条，分子量在０．３ － ３ｋｂ 之间，其中５ 种引物ＯＰＪ－ １ ，７ ，９，１１ ，１２ 扩增出差异片段１２ 条。结果表明，低能氮离子注入甜菊种子可引起体内基因组ＤＮＡ 发生突变；ＲＡＰＤ 技术是检测基因组ＤＮＡ 发生诱变的一种简便、有效方法。本文同时探讨了离子强度和Ｔａｇ ＤＮＡ 聚合酶用量对甜菊ＲＡＰＤ 分析结果的影响，以及氮离子注入诱变效应的可能机制。

广东野百合DNA提取和RAPD条件的优化

以野百合(Lilium brownii)新鲜叶片、硅胶干燥叶片及鳞片为材料,研究了DNA的提取方法,并对影响随机扩增多态DNA(RAPD)反应的各因素进行了优化。建立了野百合RAPD的优化反应体系及程序,即在20μl反应体系中,含20 ng模板DNA,2.0 mmol/L Mg2+、0.2 mmol/L dNTPs、1.5 U Taq DNA聚合酶、0.3μmol/L随机引物S1519;扩增程序为:94℃预变性5 min,然后94℃30 s,38℃50 s,72℃1 min,35个循环,最后72℃延伸10 min,4℃保存。

西伯利亚蝗基因组DNA提取及RAPD分析条件的优化

以西伯利亚蝗Gomphocerus sibiricus(L.)为研究材料,利用改良的SDS法提取高质量的DNA,分别测试了dNTP浓度、镁离子浓度、TaqDNA聚合酶用量、模板DNA的量等因素对反应结果的影响。通过各因子的组合比较,建立了西伯利亚蝗RAPD优化体系:25μLPCR反应体系,10×buffer2·5μL;dNTP0·24mmol/L;MgCl22·0mmol/L;Taq DNA聚合酶1U;DNA模板45ng;引物30ng。扩增程序为:94℃预变性1min45s、94℃变性30s、35℃退火1min30s、72℃延伸2min,45个循环、72℃延伸10min。结果表明,利用优化的反应条件进行西伯利亚蝗基因组DNA分析,实验有着良好的重复性和稳定性。

辣椒细胞质雄性不育系与其保持系线粒体DNA的RAPD分析

以辣椒细胞质雄性不育系21A及其相应的保持系21B为试材，从苗龄10～15d的黄化苗中分离纯化线粒体并提取线粒体DNA。线粒体负染后电镜观察发现，分离纯化的线粒体形态完整，呈椭球形，杂质较少。电泳检测提取的线粒体DNA的琼脂糖，结果表明，线粒体DNA纯度较高，无降解现象，浓度为20～50ng／山，可用作RAPD分析。经初步筛选，100个RAPD随机引物中有8个引物表现出多态性，经再次重复筛选，获得了辣椒细胞质雄性不育系21A线粒体DNA与雄性不育性相关的RAPD标记CMSAG3430。

芸薹类蔬菜基因组DNA遗传多样性的RAPD分析

采用随机引物扩增多态性 DNA( RAPD)技术 ,对芸薹类 ( 2 n=2 0 )蔬菜作物的 3 3个品种进行了遗传多样性检测 .从 70个引物中筛选出 3 7个引物 ,共扩增出 3 53带 .其中 ,多态带有 2 60条 ,扩增片段长度大多数集中在 0 .9～ 1 .6kb之间 .不同引物的检测效率相差很大 .运用 5个引物扩增的 1 0条RAPD特征带 ,可以作为一组 DNA指纹 ,区分所有供试