Cross‐genera amplification of informative microsatellite markers from common bean and scarlet runner bean for assessment of genetic diversity in mungbean (Vigna radiata)

Mungbean (V. radiata) is an important Asiatic legume supplying inexpensive protein to a vast majority of vegetarian masses. To increase markers repertoire in mungbean, a study was conducted to analyse 384 microsatellite markers derived from common bean, scarlet runner bean and adzuki bean for their transferability and polymorphism. The results showed that 87 (24.71%) primer pairs could amplify DNA loci of 20 mungbean genotypes including one accession of V. trilobata, while 52 showed reliable banding and polymorphism. These showed different degrees of variability at each locus producing 250 alleles with the number of alleles varying from 2 to 9. The major allele frequency varied from 0.17 to 0.95, while the polymorphic information content of SSRs ranged between 0.09 and 0.86 with an average of 0.60 ± 0.16. UPGMA revealed three major clusters accommodating ~95% of the accessions while one accession of V. trilobata (‘NSB‐007’) did not group with any other genotype describing the discriminating power of informative microsatellites. This study identified a set of useful microsatellite markers to accelerate the genetic studies and breeding programme of mungbean.     

Value of permanent grassland habitats as reservoirs of Festuca pratensis Huds. and Lolium multiflorum Lam. populations for breeding and conservation

European natural and semi-natural grassland form reservoirs of genetic resources containing highly adapted and variable ecotype populations of forage plants. Variation within these reservoirs is stimulated by variation in natural and anthropogenic site-related factors. Changes in agricultural practices lead to the loss of many characteristic habitats. In order to preserve resources for breeding, targeted conservation strategies for germplasm in gene banks (ex-situ) or on site (in-situ) are needed. In order to define site-related criteria for the potential of habitats to preserve valuable resources for breeding and conservation, 38 different habitats across Switzerland were selected to collect Festuca pratensis Huds. and Lolium multiflorum Lam. ecotype populations. Phenotypic variation and population differentiation of 60 single plants were evaluated in a field experiment using 16 morpho-physiological traits. For F. pratensis, ecotype populations and cultivars were clearly separated and there was a significant correlation between diversity of morphological traits and geographic location of sampling sites. For L. multiflorum no clear separation of ecotype populations and cultivars was observed suggesting gene flow from adjacent temporary leys into permanent grassland. Several ecotype populations were superior to cultivars in important traits such as early heading or resistance to winter damage, indicating the importance of natural habitats as a reservoir of genetic resources for breeding. In conclusion, maintenance of permanent grassland in contrasting environments appears to be a promising strategy for preserving valuable genetic variation of forage grasses. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Identification of microsatellite markers linked to the blast resistance gene <Emphasis Type="Italic">Pi-1(t)</Emphasis> in rice

The present work was conducted to identify microsatellite markers linked to the rice blast resistance gene Pi-1(t) for a marker-assisted selection program. Twenty-four primer pairs corresponding to 19 microsatellite loci were selected from the Gramene database (www. gramene.org) considering their relative proximity to Pi-1(t) gene in the current rice genetic map. Progenitors and DNA bulks of resistant and susceptible families from F₃ segregating populations of a cross between the near-isogenic lines C101LAC (resistant) and C101A51 (susceptible) were used to identify polymorphic microsatellite markers associated to this gene through bulked segregant analysis. Putative molecular markers linked to the blast resistance gene Pi-1(t) were then used on the whole progeny for linkage analysis. Additionally, the diagnostic potential of the microsatellite markers associated to the resistance gene was also evaluated on 17 rice varieties planted in Latin America by amplification of the specific resistant alleles for the gene in each genotype. Comparing with greenhouse phenotypic evaluations for blast resistance, the usefulness of the highly linked microsatellite markers to identify resistant rice genotypes was evaluated. As expected, the phenotypic segregation in the F₃ generation agreed to the expected segregation ratio for a single gene model. Of the 24 microsatellite sequences tested, six resulted polymorphic and linked to the gene. Two markers (RM1233*I and RM224) mapped in the same position (0.0 cM) with the Pi-1(t) gene. Other three markers corresponding to the same genetic locus were located at 18.5 cM above the resistance gene, while another marker was positioned at 23.8 cM below the gene. Microsatellite analysis on elite rice varieties with different genetic background showed that all known sources of blast resistance included in this study carry the specific Pi-1(t) allele. Results are discussed considering the potential utility of the microsatellite markers found, for MAS in rice breeding programs aiming at developing rice varieties with durable blast resistance based on a combination of resistance genes. Centro Internactional de Agricultura Tropical (CIAT) institute where the research was carried out     

Breeding systems in Coix lacryma-jobi populations

Summary Six populations of Coix lacryma-jobi with distinct characters were studied to evaluate the presence of autonomous apomixis and pseudogamy, and to determine its implications and effects in the calculations of the rate of breeding. For this reason, two types of genetic markers were used-morphological markers and isozymatic markers.The results show that autonomous apomixis does not seem to occur in these populations. Further experiments are necessary to prove the presence of pseudogamy in one of the populations. The rate of natural outcrossing in the grain types Rosario, Rosadlay and I-47 fluctuated around 35.9% using estimates based on isozymatic markers. The rate of natural outcrossing was 37.4% in the Adlay population using morphological markers. A population composed of tall individuals, named Tailândia, which has a greater vegetative mass and fewer grains, presented a high level of natural outcrossing at 72.8%. This information is important for many reasons, such as: 1) guiding sampling strategies of genetic material, 2) in the organization and upkeep of germplasm banks, and 3) for plant breeding programs.     

Evaluating genetic variation and relationships among two bromegrass species and their hybrid using RAPD and AFLP markers

The two most widely grown bromegrass species in North America are smooth bromegrass (Bromus inermis Leyss.), a hay type grass, and meadow bromegrass (Bromus riparius Rehmann), a pasture type grass. Recently a hybrid bromegrass population between the two species has been produced as a dual-purpose hay-pasture grass. Molecular markers have the potential to improve selection procedures to enhance bromegrass breeding. The objective of this study was to use RAPD and AFLP markers to determine genetic relationships and variations among bromegrass populations. Forty-three RAPD markers from 21 primers and 83 AFLP markers from seven primer combinations were used. Both marker types were able to group the individuals into their respective populations. The relationships among the individuals within each of the populations were not similar between the two marker types. Analysis of molecular variance (AMOVA) detected greater within-population variation than among-population variation for both marker types. The highest variation was observed in the hybrid population followed by meadow and then smooth bromegrass. The inter-population distance from both markers indicated that the highest genetic distance was between meadow and smooth bromegrass and lowest between smooth and hybrid bromegrass, which reflect the breeding history of the hybrid population. This study showed that both markers are capable of differentiating bromegrass genotypes into their respective populations, detecting genetic variation and relationships of the populations. Results of this study suggest that these two markers can be used in the future to enhance the current breeding practices in bromegrass, however, AFLP markers would be the marker of choice due to the high number of polymorphic markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification and characterization of microsatellites in eggplant

The potential of microsatellite markers for use in genetic studies in eggplant, Solanum melongena, has been evaluated. A genomic library of eggplant was screened for GA and GT repeat motifs to isolate microsatellite clones. The frequency of each repeat motif in the eggplant genome was found to be every 3200 kb for GA repeats and every 820 kb for GT repeats. Sixty‐one per cent of GT repeats were found to directly flank AT repeats. A total of 37 polymerase chain reaction (PCR) primer pairs were designed, 23 of which amplified a single product or several products. The level of microsatellite polymorphism was evaluated by using S. melongena lines and related Solanum species. Two to six alleles per primer pair were displayed in the S. melongena lines and two to 13 alleles were displayed in the Solanum relatives. Seven microsatellites showed polymorphism between parental lines of the mapping population and segregated in a codominant Mendelian manner. These microsatellite loci were distributed throughout the linkage map.     

Conservation of microsatellite flanking sequences in different taxa of Leguminosae

To test if locus-specific microsatellite markers designed for one genus are informative when used with related genera, the conservation of microsatellite-flanking intergeneric primer binding sites was tested in the closely related tribes Vicieae and Cicereae, from the subfamily Papilionoideae of the Leguminosae family. A total of 123 sequence-tagged microsatellite sites (STMS) markers derived from chickpea were used to amplify loci in lentil (Lens) and dry pea (Pisum). The percentage of chickpea primer binding sites conserved between the three genera was 54.4%. Hybridisation of 63 selected amplified loci to the digoxigenin-labelled oligonucleotide probe (TAA)₅ showed that 69.8% of loci from dry pea and 66.6% of loci from lentil hybridised to the probe. Sequencing of amplified products from chickpea with the primer Ta176 demonstrated that one amplicon contained a microsatellite, whereas another amplicon amplified with the same particular STMS primer pair did not. Amplicons produced from lentil and pea with this primer pairs did not contain microsatellite sequences. Results obtained with Tr7, which amplified a PCR product in lentil and chickpea but not in pea, showed that microsatellite sequences were present in chickpea and absent in lentil. Similar results were obtained with Ts35, which produces amplicons in pea and chickpea; but, again, microsatellite sequences were only present in chickpea. We therefore conclude that STMS derived from chickpea could be used to detect variability between other Leguminosae genera, but it is necessary to verify whether homologous loci are revealed.     

Variation of <Emphasis Type="Italic">Wx</Emphasis> microsatellite allele,waxy allele distribution and differentiation of chloroplast DNA in a collection of Thai rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Eighty-two varieties of rice from different regions in Thailand were selected to explore the Waxy (Wx)gene diversity and indica-japonica differentiation of chloroplast DNA. A comparison of the 5 splice site in the first intron was made between glutinous and nonglutinous rice. It revealed that non-glutinous with low-amylose content and glutinous rice were characterized as the Wx^b allele based on the G-to-T base substitution, whereas non-glutinous rice with intermediate and high amylose carried the Wx^a allele. Four Wx microsatellite alleles, (CT)_n repeat, (n = 16,17,18 and 19) were found in glutinous rice. In contrast, non-glutinous rice showed five Wx microsatellite alleles (n = 11, 16, 17, 18 and 19). The (CT)₁₇ allele was prominent allele in Thai population, while the (CT)₁₁ allele was found only in intermediate and high amylose rice varieties from southern Thailand. Almost all of upland rice grown by various ethnic groups in northern Thailand were characterized as japonica type based on their having the PstI-12 fragment in their cpDNA, whereas most of rainfed lowland varieties from other regions of Thailand were indica. This exploration of DNA-based genetic markers is important, as it enhances our ability to describe and manipulate sources of genetic variation for rice breeding programs.     

Changes in genetic diversity during seven cycles of recurrent selection for grainyield in oat, Avena sativa L.

Variability for desirable alleles within elite breeding populations is a requirement for long-term genetic improvement. Changes in genetic diversity were evaluated in a recurrent selection programme for enhanced grain yield in oat, Avena sativa L., using coefficient of parentage (r_p) and restriction fragment length polymorphism (RFLP) based genetic similarity (S) estimates. Both r_p and s increased during seven cycles of recurrent selection, indicating a reduction in the level of genetic diversity within the population. The relationship between genetic diversity and agronomic performance was also examined. Genetic similarity measures were poor predictors of the near-homozygous progeny performance and general and specific combining ability effects for three agronomic traits in the recurrent selection programme. In addition, there was no apparent trend for preferential selection of progeny from either more similar or more diverse parents within a given cycle of selection. The overall reduction in genetic diversity in this population has not affected selection response for grain yield. Variability remaining at important loci or de novo variation are possible explanations for the continued selection progress.     

Quantitative trait loci analysis in Theobroma cacao using molecular markers. Inheritance of polygenic resistance to Phytophthora palmivora in two related cacao populations

Two related segregating populations of Theobroma cacao L. were analysed for their resistance to Phytophthora palmivora. The first F₁ population was obtained by crossing two susceptible cacao clones of Catongo (a highly homozygous genotype) and Pound 12(a highly heterozygous genotype) and the second population was obtained by backcrossing a single F₁ tree with Catongo. The genetic maps obtained for each population were compared. The F₁ map includes 162 loci and the backcross has 140 loci. The two maps, F₁ and BC₁, exhibit high co-linear loci organization covering respectively, 772 and 944 cM.Phytophthora resistance was assessed by measuring the size increase of a lesion at five (DL5)and ten days (DL10) after pod inoculation. Six different QTL were detected in the F₁ and BC₁ populations. One QTL was found in both populations, and appeared to be a major component of disease resistance, and explaining nearly 48% of the phenotypic variance in the F₁ population. The absence of some yield QTL detection in the BC₁ in comparison with the F₁ population is due to the lack of transmission of the favouring alleles for these QTL from the single F₁ tree used for the backcross. The phenotypic variance explained by the action of the quantitative trait alleles indicated that genetic factors of both major and minor effects were involved in the control of the character studied. QTL conferring increased resistance to Phytophthorawere identified in both susceptible parents, suggesting the presence of transgressive traits and the possibility of selection in cacao. Pleiotropic and epistatic effects for the QTL were also detected. Finally, the use of marker assisted selection (MAS) in cacao breeding programs is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

用于中国小豆种质资源遗传多样性分析SSR分子标记筛选及应用

以375份小豆核心种质为试验材料, 利用从小豆及其近缘种SSR引物中筛选出的13对引物进行遗传多样性分析。检测结果显示, 小豆种质资源具有丰富的遗传多样性, 共检测到133个等位变异, 每对SSR引物检测到等位变异4~19个, 平均10.23个, 国内各省多态信息含量(PIC)平均为0.561, 多态位点比例(P)平均为93.523%。聚类结果表明, 小豆资源遗传关系与生态分区间有明显的联系, 且东北地区资源与中南部资源遗传关系较近。湖北、安徽、陕西3省资源的PIC较高, 且基本位于主坐标三维图的中心区域, 推断湖北、安徽、陕西是中国栽培小豆的起源地或多样性中心。该结果有助于更好地对小豆种质资源进行收集、保护和利用。     

Habitat and management affect genetic structure of Festuca pratensis but not Lolium multiflorum ecotype populations

Genetic diversity present in permanent grassland may be valuable for broadening gene pools in breeding programmes and for conservation of genetic resources. However, little is known about the amount of genetic diversity present at specific habitats and about site‐related factors affecting it. To identify valuable habitats, genetic diversity of 12 ecotype populations and four reference cultivars of both Festuca pratensis Huds. and Lolium multiflorum Lam. was analysed using Simple sequence repeat markers (SSR). Analysis of molecular variance revealed a larger within population variation for L. multiflorum (97.1%) than for F. pratensis (92.6%). F. pratensis ecotype populations were clearly separated from cultivars and formed three distinct subclusters according to the geographic regions they were sampled from. Differences between L. multiflorum ecotype populations and cultivars were small and no grouping of populations was observed. Thus, only F. pratensis ecotype populations were structured and habitat as well as management had a slight influence on genetic structure. This information may allow the design of individual strategies for targeted utilization of genetic resources in plant breeding programmes.     

Recurrent selection of cocoa populations in Côte d'Ivoire: comparative genetic diversity between the first and second cycles

In Côte d'Ivoire, the cocoa breeding programme has been based on the creation of hybrids between different genetic groups. From 1990 onward, a reciprocal recurrent selection programme has been set up with the purpose of improving simultaneously the characteristics of the two main genetic groups: Upper Amazon Forastero (UA) and a mixture of Lower Amazon Forastero (LA) and Trinitario (T). Based on data obtained from 12 microsatellite primers, the genetic diversity and genetic distances of the parental populations used in the first and second selection cycles are presented. The results revealed that the diversity of populations UA0 and UA1 on the one hand and (LA+T)0 and (LA+T)1 on the other is similar. The genetic distances were small between the parental populations used for the first and second cycles. Genetic diversity was greater in the UA group than in the LA+T group. The number of rare and of private alleles was reduced for both genetic groups, as well as the number of the frequent alleles in the LA+T group.     

A microsatellite sequence closely linked to the Waxy gene of Oryza sativa

Summary A polymorphic microsatellite locus has been located closely linked to the Waxy gene of rice. Primers were designed to allow detection of the microsatellite by utilising the polymerase chain reaction. In screen of 13 commercial rice varieties, four different alleles were found, demonstrating the potential of this marker in commercial rice breeding for starch quality.Abbreviations RFLP Restriction Fragment Length Polymorphism - PCR Polymerase Chain Reaxtion     

Polymorphic microsatellite marker set for <Emphasis Type="Italic">Carica papaya</Emphasis> L. and its use in molecular-assisted selection

Conventional methods for the selection of papaya pure lines are time-consuming. Procedures involving the use of molecular markers and the indirect selection of homozygous plants can reduce this time considerably. The objective of this study was to evaluate the informativeness of a microsatellite marker set when used in marker-assisted selection (MAS) for the development of new papaya lines. Eighty-three lines originating from two segregating F₃ populations and from papaya germplasm were used for the molecular analysis of 27 microsatellite primers. Twenty polymorphic microsatellite primers were identified, allowing the identification of 86 alleles, with an average of 3.18 alleles per primer. The observed heterozygosity values were low for both the markers (0.00–0.29) and the individual lines (0.00–0.35). The inbreeding coefficient (f) ranged from 0.634 to 1.00. Eleven lines with f = 1.00 and 18 lines with f varying from 0.953 to 0.961 were identified. In addition, papaya lines showed high genetic diversity, which will certainly contribute to the development of new varieties. Our results show that the use of microsatellites in MAS is a quick and effective procedure for the development of papaya lines.     

Characterisation of Cannabis accessions with regard to cannabinoid content in relation to other plant characters

Summary Ninety seven Cannabis accessions were evaluated for cannabinoid content and non-chemical plant characters. Variation within populations for cannabinoid content, and consistency of chemical characters at the population level were investigated. The relationship between chemical and other plant characters was very limited. Leaflet width and phenological data can be used for a rough prediction of the chemical phenotype on a population level. Various combinations of cannabinoid content and other economic plant characters were observed, thus a breeding programme will not be hampered by strict linkage. For a selection programme a direct analysis of cannabinoids will be inevitable.     

Grasses and <Emphasis Type="Italic">Neotyphodium</Emphasis> endophytes: co-adaptation and adaptive breeding

Neotyphodium endophytes profoundly affect the biology of their host grasses, including the important forage species perennial ryegrass, tall fescue and meadow fescue. The endophytic association is finely balanced, involving adaptation in each associate. The evidence for co-adaptation is discussed. The presence of endophyte, and the difference between endophyte strains, can affect some plant properties more than genetic variation intrinsic to the host. Conversely, host genotype exerts significant influence on the activity of the endophyte. Control of endophyte status is essential in any grass breeding programme, and developing knowledge of endophyte and its role in a herbage association offers new opportunities to grass breeders.     

Isozyme variation in Helianthus petiolaris and sunflower,H. annuus

Summary Helianthus petiolaris is a wild species used in genetic improvement of sunflower, as a donor of cytoplasmic male sterility and of genetic resistance to diseases. Isozyme variation for ADH, ACP, EST, GDH, LAP, PGI, PGD and SKDH in this species was studied using starch gel electrophoresis. The patterns thus obtained were compared with zymograms of inbred lines, hybrids and open pollinated varieties of H. annuus. The same alleles for EST and SKDH isozymes were found in both species, while ACP showed an allele that has not been found in sunflower. The rest of the isozyme systems showed both common alleles and characteristic ones for each species. ACP, GDH and PGD were monomorphic in H. petiolaris, while ADH and LAP were monomorphic in H. annuus. The isozyme markers obtained here could be useful in breeding programs involving interspecific crosses, and studies on introgression and on genetic variation in other populations of this wild species.     

Genetic variation for seed dormancy in Cuphea laminuligera and Cuphea lanceolata

Summary Various Cuphea species are adapted to temperate climates and characterized by caprylic, capric, and lauric acid-rich seed oils. These fatty acids are solely commercially available from palm kernel and coconut oils; thus, there is considerable interest in Cuphea as an alternative source of these fatty acids. Cuphea is a genus of undomesticated species characterized by seed dormancy and seed shedding. Our goal is to eliminate these domestication barriers in a few species with agronomic potential. Our objective was to survey existing Cuphea laminuligera and C. lanceolata germplasm for genetic variation for seed dormancy (germination percentage after a certain seed storage period). Recurrent half-sib family selection was practiced in various open-pollinated and synthetic populations. Half-sib family seed were evaluated for germination percentages at 26° C after various seed storage periods. There was significant genetic variation among families in every population. Additive genetic variance, half-sib family-mean heritability, and expected selection response magnitudes depended on the length of the seed storage period within a given population. Estimates of these parameters were significantly greater for longer seed storage periods within a population. Heritability estimates ranged from 0.00 to 0.79. Expected half-sib family selection responses ranged from 0.0 to 23.9%. Our study demonstrated there is substantial genetic variation for seed dormancy in C. laminuligera and C. lanceolata.     

利用微卫星标记分析建鲤种质资源的研究

为了评估和合理利用建鲤种质资源。从建鲤繁育群体中随机选取185尾鱼,用20个微卫星位点进行遗传多样性分析。结果表明：20个微卫星位点在该群体中所检测到的等位基因片段长度在114~316 bp,共检测出156个等位基因和402种基因型,各位点等位基因数为5~13个,平均7.8个,基因型数10~44种,平均20.1种;各位点观察杂合度(Ho)、期望杂合度(He)分别在0.346~0.978和0.619~0.880之间,平均分别为0.6434和0.757;所检测的20个位点多态信息含量(PIC)在0.552~0.868之间,平均为0.7253,都属于高度多态位点(PIC>0.5)。实验结果表明,该建鲤繁育群体多态信息含量丰富,遗传多样性水平较高,具有较大的选育潜力。群体内平均固定系数(FIS)为0.1479,说明该建鲤群体存在杂合子缺失现象。根据个体间的遗传距离构建的聚类图可以清楚地显示每个个体之间的遗传差异,这可为建鲤保种和繁殖配组提供依据,避免近交现象。