AFLP Analysis of Genetic Diversity in Indian Soybean [Glycine max (L.) Merr.] Varieties

AFLP technique was used to assess genetic diversity in 72 soybean varieties under cultivation in India. Selected 12 AFLP primer pairs produced 1319 products of which 1257 were polymorphic (95%). Wide variations were observed for the number of amplification products, percent polymorphism and average polymorphism information content (PIC). The Jaccard's similarity indices (J) based on the AFLP profiles of the 72 soybean varieties were subjected to UPGMA cluster analysis. The dendrogram generated revealed four major clusters, which were strongly supported by the high bootstrap values obtained from analyses of 1000 bootstrap samples. In addition, the Mantel's test for cophenetic correlation with r = 0.955 indicated very good fit of the varieties to a group in the cluster analysis. Some correspondence between the clustering pattern and the pedigree, place of release or target area of the variety was observed. Overall moderately high genetic diversity was observed which appears to be due to the higher genetic diversity prevalent in 12 of the varieties included in three diverse clusters and was indicative of the need to include more diverse germplasm in the soybean improvement programs.     

Analysis of Genetic Diversity and Relationships in Waxy Rice (Oryza sativa L.) using AFLP and ISSR Markers

Opaque endosperm is the main phenotypic indicator for waxy rice, but other phenotypic and genotypic variation among waxy rice accessions has largely been ignored. Previous studies showed that wide diversity in starch physiochemical properties exists in both indica and japonica waxy rices, especially for starch gelatinization temperature (GT) which could be divided into a high- and a low-GT group. In the present study, amplified fragment length polymorphism (AFLP) and inter-simple sequence repeat (ISSR) molecular markers were employed to examine genetic diversity and relationships of 56 waxy rice accessions. A total of 358 AFLP fragments were amplified with five primer combinations, showing a high level of polymorphism (78.3%). A total of 190 ISSR bands were generated with a single primer and a primer pair, showing a very high level of polymorphism (92.2%). The genetic distance matrices obtained from the two sets of markers were significantly correlated (r = 0.731, P = 0.004). The dendrogram generated with combined AFLP and ISSR markers could clearly differentiate the indica and japonica groups. Newly released varieties and breeding lines within each subspecies tended to be clustered together, whereas landraces were more distantly placed in the dendrogram. Only one AFLP band was found specific to the indica type, while no specific bands were found for starch GT. The implications for the conservation and breeding of waxy rice are discussed.     

AFLP analysis in pigeonpea (<Emphasis Type="Italic">Cajanus cajan</Emphasis> (L.) Millsp.) revealed close relationship of cultivated genotypes with some of its wild relatives

A total of 561 amplified fragment length polymorphism (AFLP) loci were generated and used to study the genetic diversity of wild and cultivated genotypes of pigeonpea. Out of 561 marker loci, 558 were polymorphic with an average of 76.12 bands. Analysis of molecular variation (AMOVA) revealed significant strong population structure when genotypes were structured according to continent of origin (F_ST = 0.22) also when structured into cultivated and wild genotypes (F_ST = 0.16). Maximum polymorphic loci were observed in cultivated species C. cajan (352) which is due to more number of genotypes used while, a minimum number of 45 polymorphic bands were obtained in C. acutifolius. Highest (0.291) average gene diversity was recorded in species C. mollis and lowest (0.079) average gene diversity was recorded in C. acutifolius. The 33 genotypes grouped into 14 clusters at 26% Jaccard’s similarity coefficients. Clustering analysis revealed most cultivated genotypes grouped into one major cluster while, the wild genotypes grouped into many clusters at 26% Jaccard’s similarity revealing greater diversity within wild species as compared to cultivated genotypes. From among the various cultivated genotypes studied, four genotypes BRG 3, ICP 7035, TTB 7 and ICP 8863 were molecularly and morphologically diverse and were used as parental genotypes to study nature of inheritance and to identify markers linked to sterility mosaic disease.     

Genetic diversity analysis of hulless barley from Shangri-la region revealed by SSR and AFLP markers

For adding the hulless barley resources of Shangri-la region to the global barley resource library, a basic work was done by us to assess their genetic diversity of this region. The genetic diversity of 60 hulless barley samples collected from three counties in Shangri-la region of Yunnan Province, were studied using SSR (simple sequence repeats) and AFLP (amplified fragment length polymorphism) markers. A total of 70 alleles were detected for 19 pairs of SSR primers, and 525 band containing 464 polymorphic bands were revealed for 5 pairs of AFLP primers. The value of polymorphism information content (PIC) ranged from 0.03 to 0.86 for SSR primers. The total numbers of alleles were 51, 55, 43 in three populations and the polymorphic bands were 188, 205 and 141. The genetic distances and genetic identity among the three populations showed their close relationship. The gene diversity among populations relative to the total population diversity (Gst) was 0.13 for SSR markers and 0.02 for AFLP markers and indicated that just 13 and 2% variations were among populations, respectively. The UPGMA cluster analysis revealed that all of the samples grouped randomly rather than clustered into distinct groups corresponding to their populations, row types and spring/fall types. We concluded that there was high genetic diversity in the population of Shangri-la region and the formation of diversity was related to complex environment and inhabitants’ traditional practices.     

Analysis of genetic diversity in farmers’ rice varieties in Sierra Leone using morphological and AFLP® markers

The objective of the current research was to investigate the status of rice genetic resources in post-war Sierra Leone using both morphological and amplified fragment length polymorphism (AFLP) data. Specifically, we aimed at investigating farmers’ rice genetic resources for homogeneity and differentiation, and at examining the genetic identity of similarly named varieties, including varieties within Sierra Leone, and between Sierra Leone and Guinea. This research was also motivated by the assumption that genetic erosion might have occurred as a result of the civil war. To determine the level of diversity and genetic relationships among farmers’ varieties of rice recently collected in Sierra Leone, two methods were used using subsets of the collected samples: (1) Using morphological data, 74 samples of 29 different varieties were analysed to investigate the relationship between (a) varieties grown in two districts in Sierra Leone, and (b) the two main cultivated species, Oryza sativa and Oryza glaberrima. A dendrogram largely clustered the varieties according to region and to the species to which the varieties belonged. (2) Using AFLP data, three separate investigations were conducted: (a) 33 samples of 10 varieties were investigated to evaluate diversity within and between varieties. The results indicated that the rice varieties possess different levels of intra-variety variation, whereas inter-variety diversity was high enough to distinguish one variety from the other. In particular, an AMOVA analysis revealed that 38 % of the total variation occurred within varieties, and 62 % between varieties. (b) 37 samples of 18 different varieties were investigated to determine the consistency of naming of varieties by farmers. The results showed that there was consistency in the naming by farmers of traditional varieties, but inconsistency in the naming of newly acquired varieties and cultivars. (c) 12 samples were investigated to check the identity of varieties carrying identical names collected in two separate regions, Sierra Leone and the neighbouring country of Guinea. The results indicated no close genetic relationships between the varieties found in Sierra Leone and Guinea despite similarities in the names given to these varieties by farmers, indicating the influence of different cultivation practices in the two countries.     

利用MSAP分析18个芥蓝齐口期的表观遗传多样性

本研究利用MSAP检测18个芥蓝齐口期DNA甲基化水平,分析了表观遗传多样性,探讨DNA甲基化模式对齐口期的影响。结果表明,18个芥蓝齐口期平均为50d,叶片数平均为10片,齐口期和叶片数不相关（相关系数为0.296）;变异系数分别为21%和18%;遗传距离分布在0~40,平均值为12.2276,在10.62处分为3类。MSAP分析表明,5对引物组合扩增得到432条多态性条带,201条片段表现出多态性,多态性比率为47%;Nei遗传距离分布在0.004~0.467,平均值为0.0958,表明遗传多样性水平较低;在0.04处分为3类。Mantel测验表明两种分析的遗传距离相关系数为-0.1366,显示齐口期、叶片数与DNA甲基化多态性没有相关性。DNA甲基化模式分析表明,非甲基化片段为110条,甲基化多态性片段为322条,分为3种带型,类型一为非甲基化带型（110条）,类型二为甲基化带型（110条）,类型三为半甲基化带型（152条）,非甲基化片段和半甲基化片段在不同品种之间呈现多态性,甲基化片段在不同品种之间呈现多态性与单态性相差不大,显示MSAP多态性主要来源于非甲基化和半甲基化片段,芥蓝甲基化模式以半甲基化为主。本文推测DNA甲基化水平降低参与芥蓝齐口期调控,MSAP分析既可用于基因组结构研究,又可用于基因组水平上性状的功能研究。     

Rhizobia nodulating African Acacia spp. and Sesbania sesban trees in southern Ethiopian soils are metabolically and genomically diverse

The diversity of 110 rhizobial strains isolated from Acacia abyssinica, A. seyal, A. tortilis, Faidherbia albida, Sesbania sesban, Phaseolus vulgaris, and Vigna unguiculata grown in soils across diverse agro-ecological zones in southern Ethiopia was assessed using the Biolog™ system and amplified fragment length polymorphism (AFLP) fingerprinting technique. By cluster analysis of the metabolic and genomic fingerprints, the test strains were grouped into 13 Biolog and 11 AFLP clusters. Twenty-two strains in the Biolog method and 15 strains in the AFLP analysis were linked to eight and four reference species, respectively, out of the 28 included in the study. Most of the test strains (more than 80% of 110) were not related to any of the reference species by both methods. Forty-six test strains (42% of 110) were grouped into seven corresponding Biolog and AFLP clusters, suggesting that these groups represented the same strains, or in some cases clonal descendants of the same organisms. In contrast to the strains from S. sesban, isolates from Acacia spp. were represented in several Biolog and AFLP clusters indicating the promiscuous nature of the latter and widespread occurrence of compatible rhizobia in most of the soil sampling locations. The results showed that indigenous rhizobia nodulating native woody species in Ethiopian soils constituted metabolically and genomically diverse groups that are not linked to reference species.     

Analysis of genetic diversity in a sweet potato (Ipomoea batatas L.) germplasm collection from Tanzania as revealed by AFLP

Sweet potato (Ipomoea batatas L.) is the fifth most important crop in the developing countries after rice, wheat, maize and cassava. The amplified fragment length polymorphism (AFLP) method was used to study the genetic diversity and relationships of sweet potato accessions in the germplasm collection of Sokoine University of Agriculture, Morogoro and Sugarcane Research Institute, Kibaha, Tanzania. AFLP analysis of 97 sweet potato accessions using ten primer combinations gave a total of 202 clear polymorphic bands. Each one of the 97 sweet potato accessions could be distinguished based on these primer combinations. Estimates of genetic similarities were obtained by the Dice coefficient, and a final dendrogram was constructed with the un-weight pair-group method using arithmetic average. AFLP-based genetic similarity varied from 0.388 to 0.941, with a mean of 0.709. Cluster analysis using genetic similarity divided the accessions into two main groups suggesting that there are genetic relationships among the accessions. Principal Coordinate analysis confirmed the pattern of the cluster analysis. Analysis of molecular variance revealed greater variation within regions (96.19%) than among regions (3.81%). The results from the AFLP analysis revealed a relatively low genetic diversity among the germplasm accessions and the genetic distances between regions were low. A maximally diverse subset of 13 accessions capturing 97% of the molecular markers diversity was identified. We were able to detect duplicates accessions in the germplasm collection using the highly polymorphic markers obtained by AFLP, which were found to be an efficient tool to characterize the genetic diversity and relationships of sweet potato accessions in the germplasm collection in Tanzania.     

Genetic Analysis of Egyptian Date (<Emphasis Type="Italic">Phoenix dactylifera</Emphasis> L.) Accessions Using AFLP Markers

Forty-seven samples of date palm (Phoenix dactylifera L.) collected from eight locations in Egypt were studied using four sets of amplified fragment length polymorphism (AFLP) markers with near infrared fluorescence labeled primers. These samples belonged to 21 named accessions and 9 of unknown pedigrees. A total of 350 bands were scored and 233 (66.6%) were polymorphic. Twenty-seven Egyptian accessions and ‘Medjool’and ‘Deglet Noor’accessions from California could beclassified into the major cluster. This major cluster may represent a major group of date palm germplasm in North Africa. There were four other clusters, each containing one or two accessions. The variety ‘Halawy’and one accession of unknown provenance were most likely from hybridization between two clusters. Six groups of accessions of which had the same names, revealed similar but not identical AFLP profiles suggesting these accessions might derive from seedlings rather thanthrough clonal offshoot propagation.     

紫茎泽兰群体遗传多样性及遗传结构的AFLP分析

紫茎泽兰是我国最为严重的入侵物种之一。本研究以中国境内的紫茎泽兰为材料，用AFLP方法研究了其群体遗传多样性及遗传分化。用筛选出的3对荧光引物，对5个地区62个群体进行AFLP分析，共扩增出490条带，其中多态性带328条，群体的遗传多样性较高(P = 66.9%，H = 0.171，I = 0.268)，主要存在于群体内，群体间的遗传分化系数Fst为0.287。AMOVA分析表明，在地区水平紫茎泽兰的遗传分化主要存地区内，群体内遗传分化占 70.25%，群体间遗传分化占21.71%，地区间的遗传分化仅占8.04%。UPGMA聚类分析结果表明，62个地理群体主要分为四大类群，并且有明显的地缘关系。Sigmaplot分析表明海拔是影响紫茎泽兰遗传多样性主要地理因子。Mantel检验表明遗传距离与地理距离成正相关(r = 0.34)，但也有例外。由此推断风媒传播可能是紫茎泽兰的主要传播方式，水媒传播可能是紫茎泽兰的另一主要传播途径；随着紫茎泽向东、向北扩散，新入侵地区的遗传多样性逐步降低。     

AFLP Fingerprinting in Pigeonpea (Cajanus cajan (L.) Millsp.) and its Wild Relatives

Detection of DNA polymorphism in cultivated pigeonpea (Cajanus cajan) and two of its wild relatives Cajanus volubilis and Rhynchosia bracteata is reported here for the first time using amplified fragment length polymorphism (AFLP) fingerprinting. For this purpose, two EcoRI (three selective nucleotides) and 14 MseI (three selective nucleotides) primers were used. The two wild species shared only 7.15% bands with the pigeonpea cultivars, whereas 86.71% common bands were seen among cultivars. Similarly, 62.08% bands were polymorphic between C. volubilis and pigeonpea cultivars in comparison to 63.33% polymorphic bands between R. bracteata and pigeonpea cultivars, and 13.28% polymorphic bands among pigeonpea cultivars. The cluster analysis revealed low polymorphism among pigeonpea cultivars and very high polymorphism between cultivated pigeonpea and its wild relatives. The AFLP analysis also indicated that only one primer combination (EcoRI + ACT and MseI + CTG), at the most any four primer pair combinations, are sufficient for obtaining reliable estimation of genetic diversity in closely related cultivars like pigeonpea material analyzed herein. AFLP analysis may prove to be a useful tool for molecular characterization of pigeonpea cultivars and its wild relatives and for possible use in genome mapping.     

47份水稻品种资源的ISSR遗传多样性分析

为研究广东省惠州市种植的常规水稻品种的遗传多样性,本实验利用ISSR标记对47份水稻品种资源进行遗传多样性检测。从49条引物中筛选出5条重复性好,条带清晰的引物进行PCR扩增,共扩增出53条带,每个引物可以扩增出9～13条带,平均为10．6条,其中47条具有多态性,比率为88．7％。不同水稻品种间遗传相似系数变幅为0．319～0．936,平均达0．691,说明ISSR标记能够揭示材料间较高的遗传多样性。通过聚类,从分子水平对水稻品种资源的遗传关系进行分析,并对47份水稻品种资源进行分类,ISSR标记能将47份水稻品种完全区分开,为水稻品种资源的研究利用提供参考。     

玫瑰黄链霉菌Men-myco-93-63与其突变株间的DNA多态性分析和序列特征性扩增区域(SCAR)标记

玫瑰黄链霉菌(Streptomyces roseoflavus )Men-myco-93-63是分离自马铃薯疮痂病自然衰退土壤中的一株拮抗菌。前期研究表明,该菌株及其发酵液对棉花黄萎病菌（Verticillium dahliae）等多种重要的植物病原真菌具有较强的抑制作用。利用AFLP对Men-myco-93-63及其8株抗生素生物合成阻断突变株进行了基因组DNA多态性分析。结果表明,38对AFLP引物共产生3 142条谱带,其中2 362条为多态性谱带,占总带数的75.2 %。从96对引物中筛选得到了3对引物：E-CA/M-GA,E-GA/M-AC和E-GA/M-AG,分别扩增出254、312和209 bp 3条仅在Men-myco-93-63中存在,命名为EM254、EM312和EM209。将这3个片段回收、克隆和测序,成功地将其转化为SCAR标记,可以更加方便地用于对突变株的分子检测。     

离子束介导的水稻早熟变异株系AFLP分析

用AFLP分子标记技术对N+离子束介导玉米总DNA获得的2个稳定遗传至第6代的水稻早熟变异株系C(08-5-121-6-1)和D(08-5-121-6-2)进行分析。首先从64对引物中筛选出10对多态性较好的引物,然后用这10对引物对早熟变异株系C和D分别扩增。结果显示,变异株系C、D和阴性对照水稻B(豫粳6号)与阳性对照玉米A(郑单14)之间的AFLP扩增图谱相似率分别为15.7%、16.2%和11.6%,说明变异株系与对照豫粳6号的AFLP扩增图谱存在显著差异。变异株系C和D与阴性对照豫粳6号相比,分别扩增出50条和58条差异带,其中新带分别为25条和35条,缺失带分别为19条和15条,变异株系C和D分别扩增到与阳性对照玉米相一致的目的带6条和8条,说明变异株系C和D基因组DNA与玉米基因组DNA相似性高于对照水稻。     

Exploring genetic diversity and disease response of cultivated rice accessions (<Emphasis Type="Italic">Oryza</Emphasis> spp.) against <Emphasis Type="Italic">Pyricularia oryzae</Emphasis> under rainfed upland conditions in Benin

The main goal of this study is to gain insight into the relationship between the genetic profile of cultivated rice (Oryza spp.) accessions and their resistance to rice blast. Therefore, the genetic and phenotypic variability of a set of 350 cultivated rice accessions originating from Africa (Benin, Mali and Nigeria, Ivory Coast etc.) was examined. Seventy-seven fluorescent amplified fragment polymorphism (AFLP) markers were used to gain insight into the genetic variation and to classify the germplasm collection. In addition, the rice germplasm was assessed for its resistance to blast disease caused by Pyricularia oryzae in upland field conditions. Huge differences in responses of rice accessions to P. oryzae were observed, ranging from highly susceptible to highly resistant. Twelve percent of all accessions were highly resistant to P. oryzae. Based on their AFLP marker profile these highly resistant accessions could be separated from the other accessions. Stepwise regression revealed that the best prediction of the blast resistance level was achieved with a maximum number of 13 AFLP markers. Marker CTA22 was the most important for accurate prediction of blast resistance, this marker was present in all highly resistant accessions. It can be concluded that AFLP markers are a valuable tool to screen rice accessions for their susceptibility towards blast disease and that, based on a subset of markers, it is possible to predict the resistance to rice blast.     

Genetic Diversity of Tunisian Olive Tree (Olea europaea L.) Cultivars Assessed by AFLP Markers

About 29 olive (Olea europaea L.) cultivars including oil and table olive cultivars originating from Tunisia and other Mediterranean countries, were genotyped using amplified fragment length polymorphism (AFLP) DNA markers. This technique is a rapid and efficient method for producing DNA fingerprints. Using nine AFLP primer combinations, we produced a total of 410 AFLP markers, among which 172 revealed polymorphism. The results demonstrated a high degree of polymorphism in the olive germplasm we examined with an average of 39%. These AFLP markers were analyzed to estimate genetic distances between pairs of cultivars using Jaccard’s similarity coefficient. Furthermore, cluster and principal component analyses were performed in order to identify the genetic variation patterns. Two main groups were obtained: one comprising primarily small-fruited cultivars grown mainly for oil production and the other comprising large fruited cultivars (regardless of their end-use). Our results show no evidence of clustering of olive cultivars according to their geographic origin.     

Genetic relationships among cultivated and wild <Emphasis Type="Italic">Vigna angularis</Emphasis> (Willd.) Ohwi et Ohashi and relatives from Korea based on AFLP markers

Genetic variation and relationships among members of the azuki bean complex (Vigna angularis) including wild (V. angularis var. nipponensis), weedy, and cultivated types (V. angularis var. angularis), V. nakashimae, and rice bean (V. umbellata) from Korea were examined using the Amplified fragment length polymorphism (AFLP) method. AFLP analysis of 50 accessions revealed 333 (72.1%) polymorphic fragments out of 462 fragments amplified using seven primer combinations. The number of polymorphic fragments within each species was 70 in the azuki bean complex and 41 in V. nakashimae, but there was no polymorphism in rice bean. The number of shared fragments among species ranges from 142 between the azuki bean complex and V. nakashimae to 166 between the azuki bean complex and rice bean. Within the azuki bean complex, the range of shared bands was from 231 between cultivated and weedy types to 238 between cultivated and wild types. A dendrogram generated from Jaccard’s similarity matrix was divided into three groups, which correspond to V. nakashimae, azuki bean complex, and rice bean. The relationship between azuki bean and rice bean is closer than between azuki bean and V. nakashimae. Phenetic distances averaged 0.502 between the azuki bean complex and V. nakashimae and 0.467 between the azuki bean complex and rice bean. Within the azuki bean complex, the weedy type was more closely related to wild than cultivated types. But UPGMA dendrogram of the azuki bean complex reveals that each type is not clearly isolated. These results will help to understand genetic diversity and evolutionary dynamics of Vigna in Korea.     

水稻与异科植物月见草远缘杂交研究初报

本文报道通过水稻（品种通育211）与柳叶菜科植物—-月见草进行远缘杂交,其后代出现了大量、明显的性状变异。 通过系谱选育,获得了一批在形态、生育期和多个农艺性状上有育种价值的水稻新种质。对代表性变异材料及其水稻受体亲本和供体月见草进行了AFLP分子标记分析。结果表明,月见草的花粉介入确使受体水稻材料中在DNA分子水平产生了大量的变异。文中对远缘杂交行之有效的新方法---复态导入法的特点及效应等问题进行了讨论。     

Genetic diversity of potato landraces from northwestern Argentina assessed with simple sequence repeats (SSRs)

Andean potato varieties are cultivated in the northwest of Argentina and constitute the most important staple food for the local farmers. The genetic diversity of 155 accessions conserved at the Genebank of Balcarce (INTA) was tested using four microsatellites. Three commercial potato varieties of Tuberosum group and one accession of Curtilobum group were used as outgroups. The presence of bands was scored for each microsatellite and for each accession and the data were analysed by principal coordinate analysis. The polymorphism information content was obtained for each molecular marker from banding patterns. Analysis of molecular variance was carried out with a variable number of accessions for each landrace, from different departments and sites within departments. More than one genotype was detected in the majority of the potato landraces. Some accessions within each landrace did not differentiate. AMOVA revealed that most of the genetic variation occurred among sites within departments and among local varieties. These findings are discussed considering the agricultural practices carried out in the Andean farming system.     

Assessment of Genetic Diversity in Upland Cotton (<Emphasis Type="Italic">Gossypium hirsutum</Emphasis> L.) Breeding Lines by using Amplified Fragment Length Polymorphism (AFLP) Markers and Morphological Characteristics

Genetic diversity in 24 advanced breeding lines of cotton was studied by using six amplified fragment length polymorphism (AFLP) primer-pairs and 14 morphological characters. The six selected AFLP primer-pairs generated a total of 535 amplification products, of which 460 were found to be polymorphic resulting in 85.9% polymorphism. The observed genetic distances using AFLP markers ranged from 30% to 87% with an average of 34%. A wide range of variability was observed for all the morphological traits studied. The range in taxonomic distance was from 0.60 to 2.77. Most of the lines could be clustered in two major clusters in both the analyses. But the correlation coefficient between the pair-wise distances estimated from AFLPs and the average taxonomic distances estimated from morphological characters was found to be 0.04. UPGMA cluster analysis revealed inconsistencies in the clustering patterns, as the co-phenetic correlation coefficient between the dendrograms for morphological and AFLP data indicated a poor fit for the two data types. From combined information, genotypes ‘RST-12’, ‘H 1226’, ‘P 348’, ‘KDCAKD’, ‘CISV 24’ and ‘H1222’ were found to be distinct from rest of the material.