用RAPD分子标记探讨沙拐枣属的种间关系

利用随机扩增多态性DNA（RAPD）技术分析了14种沙拐枣属（Calligonum L.)植物,通过对16个Sangon公司十聚体随机引物进行PCR扩增,3个引物能产生多态性带。对3个引物扩增产生的45条扩增产物,计算单匹配系数,应用UPGMA方法构建亲缘关系树状图。分析结果表明：（1）物种间遗传差异明显,具有丰富的遗传多样性;（2）14种沙拐枣属植物明显聚为4类,与传统的形态学分类结果基本一致。     

RAPD技术在几个苋属植物遗传分类中的应用研究

利用RAPD技术结合聚类分析,研究了亚洲和中南美洲产的苋属植物7种14个品系遗传关系。从80个10碱基长度的随机引物中筛选出8个有效引物,共扩增出103个DNA片段,平均每个引物扩增12.9个片段,其中多态性片段95个,占92．2％,依据8个有效引物扩增的DNA片段对供试材料进行了UPGMA（非加权成组算术平均法）聚类分析,结果分为4类：（1）包括A.deflexus和A.dubius各1品系的杂草苋种;（2）祖先野生种A.hybridus 1个品系;（3）包括A.hypochondriacus6品系、A.caudatus 2品系的籽粒苋种;（4）包括A.tricolor2品系和A.mangostanus 1品系的蔬菜苋种。     

Cross-priming of microsatellite loci in subfamily cyprininae (family Cyprinidae): their utility in finding markers for population genetic analysis in three Indian major carps

This study is aimed to identify polymorphic microsatellite markers and establish their potential for population genetics studies in three carp (family cyprinidae; subfamily cyprininae) species, Labeo rohita, Catla catla and Cirrhinus mrigala through use of cyprinid primers. These species have high commercial value and knowledge of genetic variation is important for management of farmed and wild populations. We tested 108 microsatellite primers from 11 species belonging to three different cyprinid subfamilies, Cyprininae, Barbinae and Leuciscinae out of which 63 primers (58.33 %) successfully amplified orthologous loci in three focal species. Forty-two loci generated from 29 primers were polymorphic in these three carp species. Sequencing of amplified product confirmed the presence of SSRs in these 42 loci and orthologous nature of the loci. To validate potential of these 42 polymorphic loci in determining the genetic variation, we analyzed 486 samples of three focal species collected from Indus, Ganges and Brahmaputra river systems. Results indicated significant genetic variation, with mean number of alleles per locus ranging from 6.80 to 14.40 and observed heterozygosity ranging from 0.50 to 0.74 in the three focal species. Highly significant (P < 0.00001) allelic homogeneity values revealed that the identified loci can be efficiently used in population genetics analysis of these carp species. Further, thirty-two loci from 19 primers were useful for genotyping in more than one species. The data from the present study was compiled with cross-species amplification data from previous results on eight species of subfamily cyprininae to compare cross-transferability of microsatellite loci. It was revealed that out of 226 heterologous loci amplified, 152 loci that originated from 77 loci exhibited polymorphism and 45 primers were of multispecies utility, common for 2–7 species.     

利用RAPD分析杜氏藻属（Dunaliella）嗜盐种间遗传多样性

本研究首次运用随机引物对杜氏藻属（Dunaliella）中6个嗜盐种的基因组DNA进行RAPD分析。筛选获得的6个有效引物共扩增出98个可重复的DNA片段,其中95条带具有多态性,多态性条带的频率为96．9％。根据系统进化树图将杜氏藻属6个种分别归于亲缘关系相对较远的2个类群中,且6个种与它们各自的生态分布联系不紧密。     

Detection of section-specific random amplified polymorphic DNA (RAPD) markers in Lilium

Random amplified polymorphic DNA (RAPD) markers were utilized for the identification of Lilium species and inter-specific hybrids. The optimum annealing temperature of the polymerase chain reaction (PCR) for the RAPD assay in Lilium was 54 °C, which is relatively higher than the temperature used for other genera reported by previous researchers. Among 76 primers used to amplify genomic DNA by PCR, 18 primers (24%) generated polymorphic DNA fragments in Lilium species and hybrids. Cultivars were also identified by RAPD markers. Some amplified fragments were unique to species of each section and to hybrids derived from these species; that is, they were the section-specific DNA markers. Sections, Sinomartagon, Leucolirion b, Leucolirion a and Archelirion could be identified by 6 section-specific markers amplified with five primers. Seven inter-section hybrids showed the section-specific bands of both parental sections, indicating that these markers would be useful for identifying the parental sections of inter-section hybrids.     

Genetic segregation of random amplified polymorphic DNA in diploid cultivated alfalfa.

Polymerase chain reaction was used, with single 10-mer primers of arbitrary sequence, to amplify random regions of genomic DNA from a diploid cultivated alfalfa backcross population. Segregation of the random amplified polymorphic DNA (RAPD) fragments was analysed to determine if RAPD markers are suitable for use as genetic markers. Of the 19 primers tested, 13 amplified a total of 37 polymorphic fragments, of which 28 (76%) segregated as dominant Mendelian traits. RAPD markers appear useful for the rapid development of genetic information in species like alfalfa where little information currently exists or is difficult to obtain.     

Application of polymerase chain reaction with specific and arbitrary primers to identification and differentiation of Leishmania parasites

Abstract Two oligonucleotide primers Lsmc1 and Lsmv1 derived from the conserved and the variable region of a major class kinetoplast DNA (kDNA) minicircle (pLURkE3) of Leishmania strain UR6 were used for the polymerase chain reaction (PCR) in order to amplify a 461-bp fragment from the kDNAs of different Leishmania species. These primers amplify the specific fragment from the kDNAs of cutaneous species only. The cutaneous species can further be distinguished by randomly amplified polymorphic DNA (RAPD) analysis of the kDNAs of these organisms using arbitrarily chosen oligonucleotides. The arbitrary primers also generate polymorphic DNA fingerprints at the genomic level with different L. donovani isolates. The results indicate that the PCR and arbitrarily primed PCR (AP-PCR) may be extremely useful approaches for identifying and distinguishing Leishmania parasites.     

用RAPD技术探讨中国枣的种下划分

利用随机扩增多态性DNA(RAPD)技术对14个枣Ziziphus jujuba 品种和1个野生种——泰山酸枣Z．spinosus的遗传变异进行了研究。从120个10-碱基随机引物中筛选出37个多态性引物用于正式扩增,共扩增出429条DNA带,其中多态性带214条,占49．88％。根据DNA扩增结果计算了品种及类型间遗传距离,并用UPGMA构建了聚类树状图。分析结果表明：龙爪枣 Z．jujuba var．tortuosa、葫芦枣 Z. jujuba var．lageniformis、无核枣 Z.jujuba var．anucleatus 等几个变种内的遗传距离大于变种间遗传距离,认为枣的变种划分是不自然的,宜并入其原变种;枣种下不宜设变种,对枣种下的众多品种,应根据品种间的遗传关系,直接划分品种群。     

利用RAPD分子标记评价仲彬草属的种间关系

利用随机扩增多态性DNA(RAPD)技术分析了14种仲彬草属Kengyilia植物的种间关系。对34个OPRON公司十聚体随机引物进行多态性筛选,20个(58．8％)能产生多态性。14个引物产生的112个DNA片断,用于计算种间Jaccard遗传相似性系数分析,在NTSYS程序中利用UPGMA构建系统发育树状图。分析结果表明：(1)14个Kengyilia物种存在较大的遗传多样性;(2)青藏高原的物种与新疆的物种 的RAPD变异极大;(3)形态相似、地理分布一致的物种有一定的亲缘关系,聚类在一起;(4)RAPD结果与形态学和细胞学等分析结果一致。RAPD分析方法将为Kengyilia系统分类提供DNA水平上丰富的资料。     

Assessment of random amplified polymorphic DNA (RAPD) for determining the origin ofYoungia koidzumiana Kitamura (compositae)

We determined the parental species ofYoungia koidzumiana (a natural interspecific hybrid) using PCR and arbitrary 10-mer primers to generate random amplified polymorphic DNA (RAPD) markers. These markers, generated by three primers, were sufficient to distinguishYoungia sonchifolia, Youngia denticulata, Youngia chelidoniifolia, andY. koidzumiana. The electrophoresis profiles of the amplified products from each of the four species were then compared. Three primers produced a total of 42 scorable markers; nine were specific markers forY. denticulata andY. chelidoni-ifolia. The length of the amplified DNA fragments ranged from 370 to 2500 b p. The three primers revealed polymorphic bands, which were indicators of the parental species ofY. koidzumiana. These bands showed a combination of specific profiles forY. denticulata andY. chelidoniifolia. Our results also were comparable to the data obtained for flowering times, floret numbers, and chromosome numbers of the four species. Therefore, we suggest thatY. koidzumiana is a hybrid betweenY. denticulata andY. chelidoniifolia}, and that RAPD markers are well suited for assessing the origins of plant species.     

三尖杉科植物RAPD分析及其系统学意义

利用随机扩增多态性ＤＮＡ（ＲＡＰＤ）技术分析了三尖杉科植物三尖杉Ｃｅｐｈａｌｏｔａｘｕｓ ｆｏｒｔｕｎｅｉ Ｈｏｏｋ．ｆ．、粗榧Ｃｅｐｈａｌｏｔａｘｕｓ ｓｉｎｅｎｓｉｓＬｉ、海南粗榧Ｃｅｐｈａｌｏｔａｘｕｓ ｈａｉｎａｎｅｎｓｉｓＬｉ和篦子三尖极Ｃｅｐｈａｌｏｔａｘｕｓ ｏｌｉｖｅｒｉＭａｓｔ。,经筛选Ｏｐｅｒｏｎ公司的４组８０个引物,其中１１４个引物的谱带清晰呈多态性。采用ＵＰＧＭＡ法对各样本     

海南主要普通野生稻的调查及利用RAPD技术进行鉴别分析

通过对海南现存的主要普通野生稻资源进行考察,报道了海南主要普通野生稻的最新生存状况;并用RAPD技术对其进行分析。从供试材料中筛选出具有多态性的RAPD引物13条,共扩增出155条带,其中多态性条带120条,多态性条带比率(PPB)值为77.42%。UPGMA聚类结果表明:海南主要普通野生稻之间遗传关系较复杂、差异较明显,并将海南主要普通野生稻资源划分为5个小类群,可以很好地将海南本地野生稻与收集保存的野生稻鉴别开来,为海南普通野生稻资源的合理利用与有效保护提供了重要依据。     

Relationships among <Emphasis Type="Italic">Leymus</Emphasis> species assessed by RAPD markers

The DNA genetic diversity of 40 accessions of genus Leymus was analyzed by random amplified polymorphic DNA (RAPD) markers. A total of 352 products were amplified by 34 10-mer arbitrary primers, among which 337 products (95.74 %) were found to be polymorphic. 5–14 polymorphic bands were amplified by each polymorphic primer, with an average of 9.91 bands. The data of 352 RAPD bands were used to generate Jaccard’s similarity coefficients and to construct a dendrogram by means of UPGMA. Great genetic diversity in genus Leymus was observed, the genetic diversity among the different species more abundant than that of the different accessions, and the different accessions in a species or the species from the same areas were clustered together.     

10种冬青属植物遗传多样性RAPD和AFLPs分析

采用RAPD和AFLP技术,对10种冬青属植物基因组进行DNA片段扩增,以研究该属种间遗传多样性.结果表明:在RAPD分析中,通过对100种10个碱基随机引物的筛选,发现11种引物能得到多态性较高扩增产物,11种引物共扩增出301条多态性条带,多态率为98.63%.在AFLP分析中,3对选择性引物组合均扩增出了丰富的多态性片段.利用RAPD和AFLP技术分析,结果按UPGMA类平均法进行聚类,聚类结果显示冬青和代茶冬青,木姜冬青和浙江冬青以及光枝刺缘冬青与毛枝三花冬青之间的亲缘关系最近.     

Monomorphic species specific DNA, detected by polymerase chain reaction with random primers

Intra- and interspecific variability of total DNA isolated from haploid megagametophytes of coniferous species was examined using polymerase chain reaction with random primers. Based on this technique, one can with certainty detect heterozygosity at gene loci carrying null alleles and thus reveal cryptic intraspecific genetic variation. Large population samples were used. Along with random amplified polymorphic DNA, i.e., widely known fragments (amplicons) polymorphic within a species, we found invariant loci lacking individual or geographic variability but differentiating species within genera and other taxa. This DNA was termed RAMD (random amplified monomorphic DNA) to distinguish it from polymorphic DNA. Our findings suggest that genetic monomorphism of species and the dual structure of the eukaryotic genome can be detected at the DNA level as was previously shown for protein gene markers.     

Interpretation of randomly amplified polymorphic DNA marker data for fingerprinting sweet potato (Ipomoea batatas L.) genotypes

In this paper we present a method for the generation of randomly amplified polymorphic DNA (RAPD) markers for sweet potato. These were applied to produce genetic fingerprints of six clonal cultivars and to estimate genetic distances between these cultivars. The level of polymorphism within the species was extremely high. From the 36-decamer random primers used, 170 fragments were amplified, of which 132 (77.6%) were polymorphic. Ten primers resulted in no detected amplification. Of the remaining 26 primers for which amplification was achieved, only one did not reveal polymorphism. Six primers used alone enabled the discrimination of all six genotypes. Pattern analysis, which employed both a classification and ordination method, enabled the grouping of cultivars and the identification of primers which gave greatest discrimination among the cultivars.     

Molecular markers for the identification of the ectomycorrhizal fungus Tuber borchii

Five species of white truffle were classified using PCR-based techniques. RAPD (random amplified polymorphic DNA) fingerprints and specific pairs of primers were used. A RAPD fragment was constant in Tuber borchii Vittad. isolates and polymorphic among the other species. Two molecular markers specific for T. borchii were developed from the sequence of the non-polymorphic RAPD fragment and from regions flanking the 5'-3' ends of a truffle gene. These markers were applied in the identification of T. borchii fruit bodies, mycelia and mycorrhizas, allowing us to monitor the development of this fungus during its entire life cycle.     

Comparative Evaluation of RAPD and AFLP Based Genetic Diversity in Brinjal (Solanum melongena)

The present investigation was carried out with an objective of evaluating genetic diversity in brinjal (Solanum melongena) using DNA markers. A total of 38 brinjal accessions including one wild-species, Solanum sisymbrifolium were characterized using random amplified polymorphic DNA (RAP D) and amplified fragment length polymorphism (AFLP) techniques. Out of 45 primers employed to generate RAPD profiles, reproducible patterns were obtained with 32 primers and 30 (93.7%) of these detected polymorphism. A total of 149 bands were obtained, out of which 108 (72.4%) were polymorphic. AFLP analysis was carried out using four primer combinations. Each of these primers was highly polymorphic. Out of 253 fragments amplified from these four primer combinations, 237 (93.6%) were polymorphic. The extent of pair-wise similarity ranged from 0.264 to 0.946 with a mean of 0.787 in RAPD, in contrast to a range of 0.103 to 0.847 with a mean of 0.434 in AFLP. The wild species clustered separately from the brinjal genotypes. In the dendrogram constructed separately using RAPD and AFLP markers, the brinjal genotypes were grouped into clusters and sub-clusters, and the varieties released by IARI remained together on both the dendrograms. All the 30 RAPD primers in combination and each of the four primer pairs in AFLP could distinguish the brinjal accessions from each other. AFLP was thus found to be more efficient than RAPD in estimation of genetic diversity and differentiation of varieties in brinjal.     

黄独遗传多样性研究

采用ISSR标记技术研究了我国14个黄独样品的遗传多样性.从55条简单重复序列引物中筛选出9条多态性引物,共扩增出70条带,其中67条多态性带,多态性比率为95.71%,平均每条引物扩增出7.8条带.黄独原变种内Nei s基因多样性(h)为0.294 9,有效等位基因数(Ne)为1.491 1,Shannon多样性指数(I)为0.444 8.种水平h为0.326 3,Ne为1.552 9,I为0.488 3.基因分化系数(Gst)为0.782 1,基因流(Nm)为0.139 3.聚类分析表明来自海南省和台湾省的样品与我国内陆的样品较早分离.据此可将来自我国内陆的样品分为5组.ISSR聚类分析基本上支持依据形态特征对黄独变种的划分.同时实验结果也表明,云南可能是黄独在我国的分化中心.     

Identification of wild and cultivated Hordeum species using two-primer RAPD fragments

Random amplified polymorphic DNAs (RAPD) analysis has been adapted to assess the degree of RAPD polymorphism within the genus Hordeum to determine if this approach can distinguish wild and cultivated species. Nineteen wild and seven cultivated accessions were evaluated using 4 random 10-mer primers. The potential of the RAPD assay was further increased by combining two primers in a single polymerase chain reaction (PCR). RAPD fragments generated by two pairs of arbitrary 10-mer primers discriminated six wild species and one cultivated species by banding profiles. The size of the amplified DNA fragments ranged from 150 to 2300 base pairs. 33 %percent of the fragments were common to both wild and cultivated species; 67% were specific to either wild or cultivated species. The average difference in fragments was less within the species than among the species. By comparing RAPD fingerprints of wild and cultivated barley, markers were identified among the set of amplified DNA fragments which could be used to distinguish wild and cultivated Hordeum species. This revised version was published online in July 2006 with corrections to the Cover Date.