Evaluation of soybean RFLP marker diversity in adapted germ plasm

Summary Soybean RFLP markers have been primarily developed and genetically mapped using wide crosses between exotic and adapted genotypes. We have screened 38 soybean lines at 128 RFLP marker loci primarily to characterize germ plasm structure but also to evaluate the utility of RFLP markers identified in unadapted populations. Of these DNA probes 70% detected RFLPs in this set of soybean lines with an average polymorphism index of 0.30. This means that only 1 out of 5 marker loci was informative between any particular pair of adapted soybean lines. The variance associated with the estimation of RFLP genetic distance (GD_R) was determined, and the value obtained suggested that the use of more than 65–90 marker loci for germ plasm surveys will add little precision. Cluster analysis and principal coordinate analysis of the GD_R matrix revealed the relative lack of diversity in adapted germ plasm. Within the cultivated lines, several lines adapted to Southern US maturity zones also appeared as a separate group. GD_R data was compared to the genetic distance estimates obtained from pedigree analysis (GD_P). These two measures were correlated with r = 0.54 for all 38 lines, but the correlation increased to r = 0.73 when only adapted lines were analyzed.     

Qualitative and quantitative characterization of RAPD variation among snap bean (Phaseolus vulgaris) genotypes

Ten snap bean (Phaseolus vulgaris) genotypes were screened for polymorphism with 400 RAPD (random amplified polymorphic DNA) primers. Polymorphic RAPDs were scored and classified into three categories based on ethidium bromide staining intensity. An average of 5.19 RAPD bands were scored per primer for the 364 primers that gave scorable amplification products. An average of 2.15 polymorphic RAPDs were detected per primer. The results show that primer screening may reduce the number of RAPD reactions required for the analysis of genetic relationships among snap-bean genotypes by over 60%. Based on the analysis of the distribution of RAPD amplification, the same number of polymorphic RAPDs were amplified from different genotypes for all RAPD band intensity levels. A comparison of RAPD band amplification frequency among genotypes for the three categories of bands classified by amplification strength revealed a measurable difference in the frequencies of RAPDs classified as faint (weakly amplifying) compared to RAPD bands classified as bold (strongly amplifying) indicating a possible scoring error due to the underscoring of faint bands. Correlation analysis showed that RAPD bands amplified by the same primer are not more closely correlated then RAPD bands amplified by different primers but are more highly correlated then expected by chance. Pairwise comparisons of RAPD bands indicate that the distribution of RAPD amplification among genotypes will be a useful criterion for establishing RAPD band identity. For the average pairwise comparison of genotypes, 50% of primers tested and 15.8% of all scored RAPDs detected polymorphism. Based on RAPD data Nei's average gene diversity at a locus was 0.158 based on all scorable RAPD bands and 0.388 if only polymorphic RAPD loci were considered. RAPD-derived 1 relationships among genotypes are reported for the ten genotypes included in this study. The data presented here demonstrate that many informative, polymorphic RAPDs can be found among snap bean cultivars. These RAPDs may be useful for the unique identification of bean varieties, the organization of bean germplasm, and applications of molecular markers to bean breeding.     

蚕豆根瘤菌接种效应研究

为发掘和利用青海冷凉地区蚕豆优良的根瘤菌种质资源,确定根瘤菌的接种效应。将分离、纯化、分子鉴定的16株蚕豆根瘤菌通过盆栽回接试验的方法进行筛选。结果表明,筛选出6株根瘤菌,它们与青海13号蚕豆共生匹配效果较好,共生固氮能力强,促进蚕豆生长效果明显。     

Genetic diversity in recent elite faba bean lines using AFLP markers

Amplified fragment length polymorphism (AFLP) markers were used to study the genetic diversity among a large set (n = 79) of inbred lines of recent elite faba bean (Vicia faba L.) cultivars with Asian, European (Northern and Southern) and North African origin. The inbred lines were analyzed using eight selected AFLP primer combinations that produced 477 polymorphic fragments. Errors when scoring repeated lanes of one pre-amplification reaction on one gel were negligible, whereas errors when scoring lanes of two individuals of the same inbred line run on different gels were markedly higher. Scoring across gels should be backed by replicates and several appropriate check entries. Based on clustering with Jaccard's similarity coefficient and Principal Coordinate Analysis, only the Asian lines were distinct as a group, the other lines showed no marked further grouping. Nevertheless, several known pedigree relationships were verified. A priori grouping of inbred lines (geographic origin and seed size) and AFLP data corroborate available information on the history of spread and cultivation of faba bean in the studied regions. Based on the diversity observed, studies especially concerning the relationship between genetic similarity based on AFLP markers and hybrid performance within the European elite germplasm have been launched.Communicated by H.F. Linskens     

Identification and potential use of a molecular marker for rust resistance in common bean

Summary The Up ₂ gene of common bean (Phaseolus Vulgaris L.) is an important source of dominant genetic resistance to the bean rust pathogen [Uromyces appendiculatus (Pers. ex Pers.) Unger var appendiculatus [syn U. Phaseoli (Reben) Wint.]. Up ₂ in combination with other rust resistance genes may be used to obtain potentially stable genetic resistance. It is difficult, however, to combine rust resistance genes effective against a single race due to epistatic interactions that frequently occur between them. A strategy that employed bulked DNA samples formed separately from the DNA of three BC₆F₂ individuals with Up ₂ and three without Up ₂ as contrasting near-isogenic lines (NILs) was used to identify random amplified polymorphic DNA fragments (RAPDs) tightly linked to the Up ₂ locus. Only 1 of 931 fragments amplified by 167 10-mer primers of arbitrary sequence in the polymerase chain reaction (PCR) was polymorphic. The RAPD marker (OA14₁₁₀₀) amplified by the 5-TCTGTGCTGG-3 primer was repeatable and its presence and absence easy to score. No recombination was observed between OA14₁₁₀₀ and the dominant Up ₂ allele within a segregating BC₆F₂ population of 84 individuals. This result suggests that OA14₁₁₀₀ and Up ₂ are tightly linked. Andean and Mesoamerican bean germ plasm, with and without the Up ₂ allele, were assayed for the presence of OA14₁₁₀₀. Apparently, the marker is of Andean origin because all Andean lines, with or without the Up ₂ allele, contained the marker, and the marker was absent in all Mesoamerican germ plasm except the lines to which Up-2 had been purposely transferred. These results suggest that OA14₁₁₀₀ will be most useful for pyramiding Up ₂ with other rust resistance genes into germ plasm of Mesoamerican origin where the marker does not traditionally exist. The use of bulked DNA samples may have concentrated resources toward the identification of RAPDs that were tightly linked to the target locus. Marker-based selection may provide an alternative to the time-consuming testcrosses required to pyramid bean rust resistance genes that exhibit epistasis.Research supported by the Michigan Agricultural Research Station and the USDA-ARS. Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the USDA and does not imply its approval to the exclusion of other products that may also be suitable     

Genetic diversity and relationship of global faba bean (<Emphasis Type="Italic">Vicia faba</Emphasis> L.) germplasm revealed by ISSR markers

Genetic diversity and relationships of 802 faba bean (Vicia faba L.) landraces and varieties from different geographical locations of China and abroad were examined using ISSR markers. A total of 212 repeatable amplified bands were generated with 11 ISSR primers, of which 209 were polymorphic. Accessions from North China showed highest genetic diversity, while accessions from central China showed low level of diversity. Chinese spring faba bean germplasm was clearly separated from Chinese winter faba bean, based on principal component analysis and UPGMA clustering analysis. Winter accessions from Zhejiang (East China), Jiangxi (East China), Sichuan (Southwest China) and Guizhou (Southwest China) were quite distinct to that from other provinces in China. Great differentiation between Chinese accessions and those from rest of the world was shown with a UPGMA dendrogram. AMOVA analyses demonstrated large variation and differentiation within and among groups of accessions from China. As a continental geographic group, accessions from Europe were genetically closer to those from North Africa. Based on ISSR data, grouping results of accessions from Asia, Europe and Africa were obviously associated with their geographical origin. The overall results indicated that the genetic relationship of faba bean germplasm was closely associated with their geographical origin and their ecological habit.     

Intra- and inter-specific variations in Lens revealed by RAPD markers

Randomly amplified polymorphic DNA (RAPD) markers were used to estimate intra- and interspecific variations in the genus Lens (lentil). Twenty cultivars of L. culinaris ssp. culinaris, including 11 microsperma (small-seeded) and nine macrosperma (large-seeded) types, and 16 wild relatives (four accessions each of L. culinaris ssp. orientalis, L. odemensis, L. nigricans and L. ervoides), were evaluated for genetic variability using a set of 40 random 10-mer primers. Fifty reproducibly scorable DNA bands were observed from ten of the primers, 90% of which were polymorphic. Genetic distances between each of the accessions were calculated from simple matching coefficients. A dendrogram showing genetic relationships between them was constructed by an unweighted pair-group method with arithmetical averages (UPGMA). This study revealed that (1) expect for L. ervoides, the level of intraspecific variation in cultivated lentil is lower than that in wild species, (2) L. culinaris ssp. orientalis is the most likely candidate for a progenitor of the cultivated species, and (3) microsperma and macrosperma cultivars were indistinguishable by the RAPD markers identified here.     

Yield stability of determinate and indeterminate dry bean cultivars

Summary Yield stability of determinate and indeterminate dry bean (Phaseolus vulgaris L.) cultivars was compared using regression of genotypic performance on environmental means. Yields of 28 dry bean cultivars differing in plant growth habit and commercial class designation were obtained from 42 Michigan performance nurseries over the 6 year period 1980 to 1985. The determinate type I large-seeded kidney and cranberry bean cultivars had below-average seed yield and large mean square deviations from regression. Lower yielding determinate small-seeded navy cultivars had low deviation mean square values, while higher yielding determinate navy cultivars had correspondingly higher mean square deviations from regression. Although seed yield of cultivars with an indeterminate growth habit was greater than determinate cultivars, prostrate type III indeterminate cultivars had deviation mean square values equivalent to those of large-seeded determinate cultivars. The erect, short vine type II indeterminate cultivars (architypes) had greater than average seed yields and minimum deviations from regression. Compared with other plant types, the architype group showed a greater yield response to more productive environments, with regression coefficient values significantly greater than unity. These results indicate that the type II growth habit offers the breeder the best opportunity of obtaining greater seed yield without incurring loss of yield stability as occurs with the type I and type III growth habits. Since the dry bean cultivars utilized in this study represent two distinct centers of domestication, the regression analysis suggests that cultivars from the predominant genetic center demonstrate more yield stability. A non-significant rank correlation coefficient between the combined and separate analyses for deviation mean square values of large-seeded cultivars implies that commercial dry bean classes should be compared separately based on center of domestication.Contributions from Michigan Bean Commission, Michigan Bean Shippers Assoc. and Michigan Agric. Exp. Sta., Michigan Agric. Exp. Sta. Journal Article No. 11986     

蚕豆种质资源清蛋白遗传多样性分析?

利用SDS-PAGE对101份蚕豆种质资源进行了清蛋白遗传多样性分析,共检测出蛋白带2625条。除共有带外,迁移率不同的谱带类型36种,其中相对分子量为92kD、75kD、62kD、40kD、34kD、17kD、13kD的谱带在检测的种质材料中出现的频率最高,分别为92.08%、90.10%、99.01%、95.05%、95.05%、98.02%、95.05%。其余29种谱带类型具有较强的多态性,多态性谱带平均为16.09条,多态性比例为44.69%。每份种质材料的清蛋白谱带数介于21～31条之间,平均25.99条;供试种质间遗传相似系数0.6111～0.9722,平均0.7122。3个地理类群内多样性指数0.9879,类群间多样性指数0.0121,表明蚕豆清蛋白的变异主要来源于类群内。聚类分析将参试种质分为6类,与以往蚕豆种质分类研究结果类似,表明清蛋白能在一定程度上反映种质间的亲缘关系。研究结果对于蚕豆蛋白质品质育种具有一定的参考价值。     

Comparative Assessment of SSR and AFLP Markers for Evaluation of Genetic Diversity and Conservation of Fig, <Emphasis Type="Italic">Ficus carica</Emphasis> L., Genetic Resources in Tunisia

This study characterises the genetic variability of fig, Ficus carica L., using simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers. It compares the efficiency and utility of the two techniques in detecting variation and establishing genetic relationships among Tunisian fig cultivars. Our results show that using both marker systems, the Tunisian fig germ plasm is characterised by having a large genetic diversity at the deoxyribonucleic acid level, as most of AFLP bands were detected and all SSR markers were polymorphic. In fact, 351 (342 polymorphic) and 57 (57 polymorphic) bands were detected using AFLP and SSR primers, respectively. SSR markers were the most polymorphic with an average polymorphic information content value of 0.94, while AFLP markers showed the highest effective multiplex ratio (56.9) and marker index (45.2). The effective marker index was recorded highest (4.19) for AFLP markers and lowest (0.70) for the SSR ones. Our results demonstrate that (1) independent as well as combined analyses of cluster analyses of SSR and AFLP fragments showed that cultivars are clustered independently from their geographical origin, horticultural classifications and tree sex; (2) the analysis of molecular variance allowed the partitioning of genetic variation within and among fig groups and showed greater variation within groups and (3) AFLP and SSR markers datasets showed positive correlation. This study suggests the SSR and AFLP markers are suitable for diversity analysis and cultivars fingerprinting. An understanding of the genetic diversity and population structure of F. carica in Tunisia can also provide insight into the conservation and management of this species.     

Comparison of RAPD and RFLP genetic markers in determining genetic similarity among Brassica oleracea L. genotypes

Genetic similarity among 45 Brassica Oleracea genotypes was compared using two molecular markers, random amplified polymorphic DNA (RAPD) and restriction fragment length polymorphisms (RFLPs). The genotypes included 37 broccolis (var. italica), five cauliflowers (var. botrytis) and three cabbages (var. capitata) which represented a wide range of commercially-available germplasm, and included open-pollinated cultivars, commercial hybrids, and inbred parents of hybrid cultivars. Fifty-six polymorphic RFLP bands and 181 polymorphic RAPD bands were generated using 15 random cDNA probes and 62 10-mer primers, respectively. The objectives were to compare RFLP and RAPD markers with regard to their (1) sampling variance, (2) rank correlations of genetic distance among sub-samples, and (3) inheritance. A bootstrap procedure was used to generate 200 random samples of size n (n=2,3,5,... 55) independently from the RAPD and RFLP data sets. The coefficient of variance (CV) was estimated for each sample. Pooled regressions of the coefficient of variance on bootstrap sample size indicated that the rate of decrease in CV with increasing sample size was the same for RFLPs and RAPDs. The rank correlation between the Nei-Li genetic similarity values for all pairs of genotypes (990) based on RFLP and RAPD data was 0.745. Differences were observed between the RFLP and RAPD dendrograms of the 45 genotypes. Overlap in the distributions of rank correlations between independent sub-samples from the RAPD data set, compared to correlations between RFLP and RAPD sub-samples, suggest that observed differences in estimation of genetic similarity between RAPDs and RFLPs is largely due to sampling error rather than due to DNA-based differences in how RAPDs and RFLPs reveal polymorphisms. A crossing algorithm was used to generate hypothetical banding patterns of hybrids based on the genotypes of the parents. The results of this study indicate that RAPDs provide a level of resolution equivalent to RFLPs for detemination of the genetic relationships among genotypes.     

Genetic diversity among elite Sorghum lines revealed by restriction fragment length polymorphisms and random amplified polymorphic DNAs

The genetic diversity of sorghum, as compared to corn, is less well characterized at the genetic and molecular levels despite its worldwide economic importance. The objectives of this study were to: (1) investigate genetic diversity for restriction fragment length polymorphism (RFLPs) and random amplified polymorphic DNAs (RAPDs) in elite sorghum lines, (2) compare similarities based on molecular markers with pedigree relationships, and (3) examine the potential of RFLPs and RAPDs for assigning sorghum lines to the A/B (sterile) and R (restorer) groups. Using four restriction enzymes, polymorphism was detected with 61% of the RFLP probes used, compared to 77% of the random primers. One hundred and sixteen (64%) probe-enzyme combinations yielded multiple-band profiles compared to 98% of the random primers. RFLP profiles generated 290 polymorphic bands compared to 177 polymorphic RAPDs. Pair-wise comparisons of polymorphic RFLPs and RAPDs were used to calculate Nei and Jaccard coefficients. These were employed to generate phenograms using UPGMA and neighborjoining clustering methods. Analysis of RFLP data with Jaccard's coefficient and neighbor-joining clustering produced the phenogram with the closest topology to the known pedigree.Contribution of the College of Agricultural Sciences, Texas Tech University, Journal No. T-4-365     

RFLP analyses of early-maturing European maize germ plasm

Summary Thirty inbred lines representing a wide range of early-maturing European elite germ plasm of maize (Zea mays L.) were assayed for RFLPs using 203 clone-enzyme combinations (106 DNA clones with restriction enzymes EcoR1 and HindIII). The genetic materials comprised 14 flint, 12 dent, and 4 lines of miscellaneous origin. Objectives were to (1) characterize the genetic diversity for RFLPs in these materials, (2) compare the level of genetic diversity found within and between the flint and the dent heterotic groups, and (3) examine the usefulness of RFLPs for assigning inbreds to heterotic groups. All but two DNA clones yielded polymorphism with at least one restriction enzyme. A total of 82 and 121 clone-enzyme combinations gave single-banded and multiple-banded RFLP patterns, respectively, with an average of 3.9 and 7.7 RFLP patterns per clone-enzyme combination across all 30 inbreds, respectively. Genetic similarity (GS) between lines, estimated from RFLP data as Dice's similarity coefficient, showed considerable variation (0.32 to 0.58) among unrelated inbreds. The mean GS for line combinations of type flint x dent (0.41) was significantly smaller than for unrelated flint lines (0.46) and dent lines (0.46), but there was considerable variation in GS estimates of individual line combinations within each group. Cluster and principal coordinate analyses based on GS values resulted in separate groupings of flint and dent lines in accordance with phylogenetic information. Positioning of lines of miscellaneous origin was generally consistent with expectations based on known breeding behavior and pedigrees. Results from this study corroborated that RFLP data can be used for assigning inbreds to heterotic groups and revealing pedigree relationships among inbreds.     

Estimation of sampling variance of molecular marker data using the bootstrap procedure

Knowledge of genetic relationships among genotypes is useful in a plant breeding program because it permits the organization of germplasm and provides for more efficient sampling. The genetic distance (GD) among genotypes can be estimated using random restriction fragment length polymorphisms (RFLPs) as molecular markers. Knowledge of the sampling variance associated with RFLP markers is needed to determine how many markers are required for a given level of precision in the estimate of GD. The sampling variance for GD among all pairs of 37 maize (Z. mays L.) inbred lines was estimated from 1202 RFLPs. The 1202 polymorphisms were generated from 251 enzyme-probe combinations (EPC). The sampling variance was used to determine how large a sample of RFLPs was required to provide a given level of precision. The coefficient of variation (CV) associated with GD has a nearly linear relationship between its expected standard deviation and mean. The magnitude of the decrease in the mean CV for GD with increasing numbers of bands was dependent upon the sampling unit; e.g., individual polymorphic bands vs EPC, and the degree of relatedness among the inbreds compared. The rate of reduction in mean CV with increasing sample size was the same regardless of the restriction enzyme used, BamHI, EcoRI or HindIII, when the bootstrap sampling units were individual polymorphic bands. In constrast, although the rate of reduction (slopes) was the same, the intercepts of the mean CVs were different when EPCs were used as the bootstrap sampling unit. This difference was due to the higher number of bands per EPC in BamHI (4.94) compared with EcoRI (4.83) and HindIII (4.63).     

Potential taxonomic use of random amplified polymorphic DNA (RAPD): a case study in Brassica

Summary The potential use of RAPDs for taxonomic studies were investigated using Brassica, Sinapis and Raphanus taxa. Principal coordinate analysis of 284 RAPD bands revealed the classical U triangle relationship between diploid and amphidiploid Brassica taxa. Raphanus sativus and S. alba were distinct from the Brassica taxa. It appears that at least ten primers with approximately 100 total bands are needed to adequately portray these relationships. Cultivars of cabbage and cauliflower were separated by RAPDs. Analysis of RAPDs from individual plants of B. carinata cv. dodola resulted in 69 RAPDs, with 91.7% monomorphic and 8.3% polymorphic bands. RAPDs appear to be useful for taxonomic studies at levels ranging from populations to species and perhaps genera.     

Host differentiation in Orobanche foetida Poir

Orobanche foetida Poir. is a parasitic plant widely distributed in the Western Mediterranean area. It typically parasitizes wild plants but has recently been described as an agricultural problem in legume crops in Tunisia. The pattern of genetic variation within and among O. foetida populations growing on chickpea and faba bean was analyzed by RAPD markers. The UPGMA cluster analysis based on Dice distance matrix showed a clear differentiation among O. foetida samples collected on chickpea and those on faba bean, suggesting a host-differentiation process. Although an AMOVA analysis revealed substantial internal variation among individuals within O. foetida populations (69.8%), there was a significant divergence between parasites on the two hosts considered (30.2%). Moreover, germination of O. foetida seeds collected on chickpea and faba bean in the presence of both host roots was studied. Germination percentages of O. foetida seeds varied depending on the host used both for collecting the seeds and evaluating the trait. According to these results, possible explanations for the origin of this new weedy parasite and the host differentiation process are discussed.     

Genetic distances revealed by morphological characters,isozymes, proteins and RAPD markers and their relationships with hybrid performance in oilseed rape (<Emphasis Type="Italic">Brassica napus</Emphasis> L.)

Genetic distances (GDs) based on morphological characters, isozymes and storage proteins, and random amplified polymorphic DNAs (RAPD) were used to predict the performance and heterosis of crosses in oilseed rape (Brassica napus L.). Six male-sterile lines carrying the widely used Shaan2A cytoplasm were crossed with five restorer lines to produce 30 F₁ hybrids. These 30 hybrids and their parents were evaluated for seven agronomically important traits and their mid-parent heterosis (MPH) at Yangling, Shaanxi province in Northwest China for 2 years. Genetic similarity among the parents based on 34 isozyme and seven protein markers was higher than that based on 136 RAPDs and/or 48 morphological markers. No significant correlation was detected among these three sets of data. Associations between the different estimates of GDs and F₁ performance for some agronomic traits were significant, but not for seed yield. In order to enhance the predicting efficiency, we selected 114 significant markers and 43 favoring markers following statistical comparison of the mean values of the yield components between the heterozygous group (where the marker is present only in one parent of each hybrid) and the homozygous group (where the marker is either present or absent in both parents of each hybrid) of the 30 hybrids. Parental GD based on total polymorphic markers (GD_total, indicating general heterozygosity), significant markers (GD_sign, indicating specific heterozygosity) and favoring markers (GD_favor, indicating favoring-marker heterozygosity) were calculated. The correlation between GD_favor or GD_sign and hybrid performance was higher than the correlation between GD_total and hybrid performance. GD_sign and GD_favor significantly correlated with plant height, seeds per silique and seed yield, but not with the MPH of the other six agronomic traits with the exception of plant height. The information obtained in this study on the genetic diversity of the parental lines does not appear to be reliable for predicting F₁ yield and heterosis.     

Genetic variability within isolates of Colletotrichum lindemuthianum belonging to race 65 from the state of Minas Gerais, Brazil

Colletotrichum lindemuthianum, the causal agent of anthracnose in the common bean (Phaseolus vulgaris), presents a wide genetic and pathogenic variability that gives rise to complications in the development of resistant bean cultivars. The aim of this study was to identify the variability within race 65 of C. lindemuthianum, the race most commonly encountered in Brazil, through randomly amplified polymorphic DNA (RAPD) and anastomosis analyses. Thirteen isolates of race 65, collected in different years and from various host cultivars located in diverse areas of the state of Minas Gerais, Brazil, were investigated. Twenty-four RAPD primers were employed and 83 polymorphic bands amplified. Genetic similarities were estimated from the Sorensen-Dice coefficient and ranged from 0.54 to 0.82. The dendrogram obtained by cluster analysis classified the isolates into 11 separate groups. For the purposes of the analysis of anastomosis, isolates were considered to be compatible when the fusion of hyphae from different isolates could be observed. The proportion of compatible reactions for each isolate was estimated and similarity estimates, based on the Russel & Rao coefficient, ranged from 0.28 to 0.85. Isolates were classified into 11 anastomosis groups, 10 of which were formed by only one isolate. Although isolates LV61, LV73 and LV58 were classified in the same anastomosis group, they were genetically distinct according to RAPD analysis. Results from both RAPD and anastomosis analyses revealed great variability within C. lindemuthianum race 65.     

Assessment of genetic variation withinBrassica campestris cultivars using amplified fragment length polymorphism and random amplification of polymorphic DNA markers

Genetic relationships were evaluated among nine cultivars ofBrassica campestris by employing random amplification of polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) markers. RAPDs generated a total of 125 bands using 13 decamer primers (an average of 9.6 bands per assay) of which nearly 80% were polymorphic. The per cent polymorphism ranged from 60–100%. AFLP, on the other hand generated a total of 319 markers, an average of 64 bands per assay. Of these, 213 were polymorphic in nature (66.8%). AFLP methodology detected polymorphism more efficiently than RAPD approach due to a greater number of loci assayed per reaction. Cultivar-specific bands were identified, for some cultivars using RAPD, and for most cultivars with AFLP. Genetic similarity matrix, based on Jaccard’s index detected coefficients ranging from 0.42 to 0.73 for RAPD, and from 0.48 to 0.925 for AFLPs indicating a wide genetic base. Cluster analyses using data generated by both RAPD and AFLP markers, clearly separated the yellow seeded, self-compatible cultivars from the brown seeded, self-incompatible cultivars although AFLP markers were able to group the cultivars more accurately. The higher genetic variation detected by AFLP in comparison to RAPD was also reflected in the topography of the phenetic dendrograms obtained. These results have been discussed in light of other studies and the relative efficiency of the marker systems for germplasm evaluation.     

Similarities and relationships among populations of the bulb onion as estimated by nuclear RFLPs

Random nuclear restriction fragment length polymorphisms (RFLPs) were used to assess similarities and relationships among open-pollinated (OP) populations of the cultivated bulb onion (Allium cepa). Seventeen OP populations and 2 inbreds of contrasting daylength response [termed by convention as long (LD) and short (SD) day], 1 shallot (A. cepa var. ascalonicum), and one cultivar of bunching onion (Allium fistulosum) were examined with 104 cDNA clones and two to four restriction enzymes. Sixty (58%) clones detected at least 1 polymorphic fragment scorable among the OP populations and were used for analyses. The average number of polymorphic fragments per polymorphic probe-enzyme combination was 1.9, reflecting that numerous monomorphic fragments were usually present. Similarities were estimated as the proportion of polymorphic fragments shared by 2 populations. Average similarity values among LD, among SD, and between LD and SD OP populations were 0.79, 0.67, and 0.68, respectively. Relationships among the OP populations were estimated by parsimony, cluster analysis of similarities using the unweighted-pair-group method (UPGMA), and multivariate analysis using principle components. Parsimony analysis generated a strict consensus tree that grouped all but 1 LD onion with unresolved relationships to the SD OP populations. The UPGMA analysis placed together the LD storage OP populations. Principal component analysis grouped all but 2 LD onions; the other OP populations were dispersed. The results suggest that LD and SD onions do not represent distinct germ plasm, but that LD storage onions represent a derived group selected for production at higher latitudes. If it is assumed that the sampled populations are representative of all onion OP populations, the lower similarities among SD OP populations indicate that their collection and maintenance in germ plasm collections is important for the preservation of genetic diversity.