Assessment of genetic diversity among and within Carthamus species using sequence-related amplified polymorphism (SRAP) markers

Due to precise evaluation of genetic diversity of Carthamus species, sixty-two genotypes consisting fifty-two from five wild (C. oxyacanthus M. Bieb, C. lanatus L., C. dentatus Vahl, C. boissieri Halácsy, C. glaucus M.B.) and ten from cultivated species (C. tinctorius L.) were selected for evaluation of the genetic diversity in Carthamus species. A total of 238 (81.2 %) polymorphic bands were detected by 12 SRAP primer combinations with an average of 22 bands per combination. Me4-Em1 and Me5-Em2 primer combinations were known as the most informative SRAP markers based on the PIC values (0.34) where they distinguished all studied Carthamus species. Cluster analysis classified all accessions into five main groups among which clusters containing cultivated individuals were distinctly separated from those containing wilds. The most and the least genetic variation based on analysis of molecular variance, were detected within (76.90 %) and among (22.84 %) groups, respectively. The obtained results suggested that C. dentatus, C. glaucus and C. boissieri species may be classified in one section including C. dentatus in one and C. glaucus and C. boissieri in another subsection. The results also revealed high genetic similarity between C. oxyacanthus and C. tinctorius despite their different morphological characteristics.     

Genetic diversity of endangered Manglietia patungensis assessed by inter simple sequence repeat and sequence-related amplified polymorphism markers

Manglietia patungensis Hu is an endangered plant native to China. Knowledge of its genetic diversity and structure would aid its conservation. This study assessed nine natural populations of M. patungensis using two methods: inter simple sequence repeat (ISSR) and sequence-related amplified polymorphism (SRAP) markers. Using 10 ISSR primer pairs, 334 bands were generated, and 10 SRAP primer pairs generated 276 bands. The percent of polymorphic bands (91.32% and 93.48%), Nei's genetic diversity (0.3448 and 0.3323), and Shannon's information index (0.5075 and 0.4935) revealed a high level of genetic diversity at the species level. Total heterozygosity was 0.3439 by ISSR and 0.3281 by SRAP. The mean heterozygosity was 0.2323 by ISSR and 0.2521 by SRAP. The coefficient of genetic differentiation among natural populations was 0.3245 by ISSR and 0.2316 by SRAP. These data indicated higher levels of genetic diversity of M. patungensis within, rather than among, populations. Estimates of gene flow among natural populations were 1.0411 and 1.0589, which implied a certain amount of gene exchange among populations. A Mantel test revealed no significant correlation between genetic and geographic distance. ISSR and SRAP markers are both effective for genetic diversity research in M. Patungensis. Based on these results, conservation of M. patungensis should be performed both in situ and ex situ.     

Genetic characterization of Iranian native Bombyx mori strains using amplified fragment length polymorphism markers

Genetic relationships within and among seven Iranian native silkworm strains was determined by DNA fingerprinting by using amplified fragment length polymorphism (AFLP) markers. In total, 189 informative AFLP markers were generated and analyzed. Estimates of Nei's gene diversity for all loci in individual strains showed a higher degree of genetic similarity within each studied strain. The highest and the least degrees of gene diversity were related to Khorasan Pink (h = 0.1804) and Baghdadi (h = 0.1412) strains, respectively. The unweighted pair-group method with arithmetic average dendrogram revealed seven strains of silkworm, Bombyx mori (L.), resolving into two major clusters. The highest degree of genetic similarity was related to Baghdadi and Harati White, and the least degree was related to Guilan Orange and Harati Yellow. The genetic similarity estimated within and among silkworms could be explained by the pedigrees, historical and geographical distribution of the strains, effective population size, inbreeding rate, selection intensity, and gene flow. This study revealed that the variability of DNA fingerprints within and among silkworm strains could provide an essential basis for breeders in planning crossbreeding strategies to produce potentially hetrotic hybrids in addition to contributing in conservation programs.     

Genetic diversity in Capsicum germplasm based on microsatellite and random amplified microsatellite polymorphism markers

A sound knowledge of the genetic diversity among germplasm is vital for strategic germplasm collection, maintenance, conservation and utilisation. Genomic simple sequence repeats (SSRs) and random amplified microsatellite polymorphism (RAMPO) markers were used to analyse diversity and relationships among 48 pepper (Capsicum spp.) genotypes originating from nine countries. These genotypes covered 4 species including 13 germplasm accessions, 30 improved lines of 4 domesticated species and 5 landraces derived from natural interspecific crosses. Out of 106 SSR markers, 25 polymorphic SSR markers (24 %) detected a total of 76 alleles (average, 3.04; range, 2–5). The average polymorphic information content (PIC) was 0.69 (range, 0.29–0.92). Seventeen RAMPO markers produced 87 polymorphic fragments with average PIC of 0.63 (range, 0.44–0.81). Dendrograms based on SSRs and RAMPOs generated two clusters. All 38 Capsicum annuum genotypes and an interspecific landrace clustered together, whereas nine non-annuum (three Capsicum frutescens, one Capsicum chinense, one Capsicum baccatum and four interspecific landraces) genotypes clustered separately. Genetic variation within non-annuum genotypes was greater than the C. annuum genotypes. Distinctness of interspecific derivative landraces grown in northeast India was validated; natural crossing between sympatric Capsicum species has been proposed as the mechanism of their origin.     

DNA fingerprinting of Leonurus cardiaca L. germplasm in Iran using amplified fragment length polymorphism and inter-retrotransposon amplified polymorphism

In the present study, the extent of inter and intra-population genetic variation was evaluated in Leonurus cardiaca accessions naturally growing in Iran by AFLP and IRAP markers. The fingerprints corresponding to AFLP and IRAP markers revealed high levels of heterozygosity, indicating that L. cardiaca is predominantly an out-crossing species. The average percentage polymorphism was detected as 58% and 90.8% on utilizing AFLP and IRAP data, respectively. Gene diversity values within populations varied 0.14 to 0.20 for AFLP and 0.12 to 0.21 for IRAP. The overall levels of genetic variation present in the L. cardiaca germplasm in Iran were finally determined by combining the AFLP and IRAP datasets to ensure wide genome coverage. The phenogram depicted that the accessions of Dargaz population were genetically distinct from other populations. Based on AFLP and IRAP analysis, it is concluded that L. cardiaca maintains high levels of genetic variation at inter and intra-population level.     

Genetic diversity and relationships within sweet kernel apricot and related Armeniaca species based on sequence-related amplified polymorphism markers

Sequence-related amplified polymorphism (SRAP) markers were employed to detect the genetic variation among 28 sweet kernel apricot and related Armeniaca accessions from China. Fifteen SRAP primer combinations were used and 252 polymorphic bands out of 286 with an average of 16.8 per primer combination were observed. The unweighted pair-group method (UPGMA) analysis demonstrated that the accessions had a similarity range from 0.55 to 0.91 with a mean of 0.7, and there were three clusters with two groups of sweet kernel apricots distinctly separated from the related Armeniaca vulgaris and Armeniaca sibirica at the similarity level of 0.58, indicating more extensive genetic variation among sweet kernel apricot. The origin of sweet kernel apricot has also been discussed. This suggested there are abundant germplasm resources of sweet kernel apricot from China for further breeding and biodiversity conservation.     

Genetic diversity and relationships among Canavalia ensiformis (L.) DC. Accessions as revealed by sequence-related amplified polymorphism markers

Canavalia ensiformis is an under-exploited legume that has been used as forage, green manure, and a cover crop. Thus far, studies of the C. ensiformis germplasm have focused on morphological traits, which cannot be used to distinguish all known accessions or to evaluate their genetic diversity precisely. In this study, sequence-related amplified polymorphism (SRAP) markers were used to assess the genetic diversity and relationships among 29 C. ensiformis accessions originating from 16 countries. In total, 274 clear bands were amplified and 144 of them (52.6%) were polymorphic. The polymorphism information content values (PIC) ranged from 0.10 to 0.43, with an average of 0.27. An analysis of molecular variance (AMOVA) revealed that the most significant variation (92.0% of the total) occurred among accessions; the remaining 8.0% was attributed to variation within accessions. A cluster analysis and principal coordinates (PCoA) analysis produced similar results, whereby the 29 C. ensiformis accessions were divided into 5 clusters, each of which was composed of different accessions with different phenotypic traits. This study provides the theoretical basis for future biodiversity studies and breeding programs.     

Genetic characterization of <Emphasis Type=Italic>Meloidogyne incognita</Emphasis> isolates from Turkey using sequence-related amplified polymorphism (SRAP)

Southern root knot nematode Meloidogyne incognita is the most widespread-species, causing serious yield losses in protected vegetables fields in the West Mediterranean region of Turkey. The knowledge of genetic variation within M. incognita is required for disease management and improvement of resistant varieties by breeding programs. In the present study, the isolates were classified into different groups based on sequence-related amplified polymorphism (SRAP) fingerprints. To our knowledge, this is the first study carried out on the characterization of M. incognita isolates using SRAP. The schematic diagram by tested primers to differentiate of M. incognita isolates was formed in discrimination of nematodes as an effective molecular tool since it is cost effective and easiness. Data presents a genetic variation on root-knot nematode species. These selected SRAP markers can be used to follow genetic structure and differentiation on M. incognita isolates in a certain region.     

Assessment of genetic diversity of cultivated and wild Iranian grape germplasm using retrotransposon-microsatellite amplified polymorphism (REMAP) markers and pomological traits

Molecular Biology Reports - Understanding the genetic diversity and relationships between genotypes is an effective step in designing effective breeding programs. Insertional polymorphisms of...     

Molecular characterization and genetic mapping of random amplified microsatellite polymorphism in barley

 This study has analyzed the molecular basis and genetic behaviour of the polymorphism generated by the amplification of barley genomic DNA with primers complementary to microsatellites. Primers anchored at the 5′ end, used alone or in combination with arbitrary sequence primers, generated random amplified microsatellite polymorphisms (RAMPs). Unanchored primers were also used as single primers in a microsatellite primed-PCR (MP-PCR). Twenty six randomly selected RAMP DNA fragments which showed polymorphism between the cultivars Steptoe and Morex were cloned and sequenced. All sequences showed the expected repeated motif at the end of the insert, with the number of repeats ranging from five to ten. Genomic sequences containing low numbers of microsatellite motifs were preferentially amplified; therefore, only a fraction of the polymorphism could be attributed to variation in the number of microsatellite motifs at the priming site. Some sequences contained either cryptic simple sequences or members of families of repeated DNA. Polymorphism at the internal cryptic simple sequences was detected by RAMP bands inherited as co-dominant markers. Four MP-PCR bands were cloned and sequenced. A number of repeats identical to the primer itself were found at each end of the insert. Two allelic bands were polymorphic for an internal microsatellite. The potential use of cloned bands as fingerprinting tools was investigated by employing them as hybridization probes in Southern blots containing digested barley DNA from a sample of cultivars. RAMP probes produced complex hybridization band patterns. MP-PCR probes produced either a highly variable single locus or low-copy number loci. Segregations for 31 RAMPs and three MP-PCR bands were studied in a population of 70 doubled-haploids from the Steptoe/Morex cross. One third of all markers were co-dominantly inherited. Markers were positioned on an RFLP map and found to be distributed in all barley chromosomes. The new markers enlarged the overall length of the map to 1408 cM. Received: 6 May 1998 / Accepted: 20 July 1998     

Molecular characterization of Fusarium head blight resistance in Wangshuibai with simple sequence repeat and amplified fragment length polymorphism markers.

Molecular mapping of Fusarium head blight (FHB) resistance quantitative trait loci (QTL) and marker-assisted selection of these QTL will aid in the development of resistant cultivars. Most reported FHB resistance QTL are from 'Sumai 3' and its derivatives. 'Wangshuibai' is a FHB-resistant landrace that originated from China and is not known to be related to 'Sumai 3'. A mapping population of 139 F(5:6) recombinant inbred lines was developed from a cross of 'Wangshuibai' and 'Wheaton'. This population was developed to map the FHB-resistant QTL in 'Wangshuibai' and was evaluated twice for Type II FHB resistance. A total of 1196 simple sequence repeat and amplified fragment length polymorphism markers were screened on this population, and four FHB resistance QTL were detected. A major QTL near the end of 3BS explained 37.3% of the phenotypic variation. Another QTL on 3BS, located close to the centromere, explained 7.4% of the phenotypic variation. Two additional QTL on 7AL and 1BL explained 9.8% and 11.9% of the phenotypic variation, respectively. The simple sequence repeat and amplified fragment length polymorphism markers closely linked to these QTL may be useful for stacking QTL from 'Wangshuibai' and other sources to develop cultivars with transgressive FHB resistance.     

Molecular characterization of sawtooth oak (Quercus acutissima) germplasm based on randomly amplified polymorphic DNA

Sawtooth oak (Quercus acutissima) is a predominant tree species in the deciduous broad-leaved forest in China. It distributes in a large landscape area and can disperse in various ecology types. Molecular study on sawtooth oak can provide valuable information about the genetic diversity level and genetic relatedness on this important tree species. Insight into the genetic structure also provides resources of a species with its current feature and future evolutionary potential. The genetic structure of sawtooth oak was investigated by randomly amplified polymorphic DNA (RAPD). Twelve RAPD markers were used to assess genetic diversity of 408 individuals from 17 provenances enveloping most of the current distribution area of sawtooth oak. A total of 66 amplification products were detected, of which 49 bands (74.24 %) were polymorphic. Nei’s gene diversity, 0.2409, indicated a relatively high level of genetic variation in sawtooth oak germplasm. Analysis of molecular variance showed that most of the genetic diversity (87 %) was allocated within provenances. A combination of UPGMA dendrogram and STRUCTURE analysis was employed to estimate the genetic relationships of sawtooth oak germplasm; interestingly, the two methods presented similar grouping pattern with few discrepancies. Results revealed that 16 out of 17 provenances were clustered into one group, while the other 1 (LQ provenance) constituted a separate cluster. The data presented in this study suggested that the RAPD method was a valuable tool for estimation of genetic diversity and genetic relatedness of sawtooth oak germplasm. The present study also gave useful implications for germplasm conservation and new cultivar development for this promising energy tree species.     

Genetic differentiation analysis of African cassava (Manihot esculenta) landraces and elite germplasm using amplified fragment length polymorphism and simple sequence repeat markers

Molecular‐marker‐aided evaluation of germplasm plays an important role in defining the genetic diversity of plant genotypes for genetic and population improvement studies. A collection of African cassava landraces and elite cultivars was analysed for genetic diversity using 20 amplified fragment length polymorphic (AFLP) DNA primer combinations and 50 simple sequence repeat (SSR) markers. Within‐population diversity estimates obtained with both markers were correlated, showing little variation in their fixation index. The amount of within‐population variation was higher for landraces as illustrated by both markers, allowing discrimination among accessions along their geographical origins, with some overlap indicating the pattern of germplasm movement between countries. Elite cultivars were grouped in most cases in agreement with their pedigree and showed a narrow genetic variation. Both SSR and AFLP markers showed some similarity in results for the landraces, although SSR provided better genetic differentiation estimates. Genetic differentiation (Fst) in the landrace population was 0.746 for SSR and 0.656 for AFLP. The molecular variance among cultivars in both populations accounted for up to 83% of the overall variation, while 17% was found within populations. Gene diversity (H_e) estimated within each population varied with an average value of 0.607 for the landraces and 0.594 for the elite lines. Analyses of SSR data using ordination techniques identified additional cluster groups not detected by AFLP and also captured maximum variation within and between both populations. Our results indicate the importance of SSR and AFLP as efficient markers for the analysis of genetic diversity and population structure in cassava. Genetic differentiation analysis of the evaluated populations provides high prospects for identifying diverse parental combinations for the development of segregating populations for genetic studies and the introgression of desirable genes from diverse sources into the existing genetic base.     

Multi-locus DNA fingerprinting and genetic diversity in jute (Corchorus spp.) based on sequence-related amplified polymorphism

Sequence-Related Amplified Polymorphism (SRAP) markers were used for genetic diversity assessment and cultivar identification among 31 cultivars of jute belonging to two cultivated species Corchours olitorius L. and C. capsularis L. Forty-three primer-pairs produced a total of 394 bands with an average of 9 bands per primer pair and 89% bands were polymorphic across the genotypes of two species. Average genetic diversity in the cultivars of C. olitorius and C. capsularis was 7.2% (range 2.8–12.3%) and 7.6% (range 2.2–13.1%), respectively. Jute cultivars JRC 698, JRC 7447, TJ 40, S19 and JRO 3690 were more diverse compared to rest of the cultivars. UPGMA cluster analysis grouped all cultivars into two clusters which were representative of C. olitorius and C. capsularis species. All the cultivars could be unequivocally differentiated from one another based on the pooled profile of 43 primer-pairs, however, 24 of 31 cultivars could be identified uniquely. The probability of chance identity of any two cultivars based on SRAP markers was very low and was 6.95?×?10^?07 and 2.23?×?10^?07 for cultivars of C. capsularis and C. olitorius, respectively. Primer-pairs EM1-ME5, EM4-ME1, EM8-ME1 and EM10-ME1 were found to be useful for genetic diversity and cultivar identification. Our results show that SRAP markers could be effectively used for genetic diversity analyses in jute. For poor genetic diversity and resulting narrow genetic base, these markers will prove to be highly useful for identifying elite germplasm in a jute breeding program.     

Molecular characterization of Lebanese olive germplasm

Lebanon, an East Mediterranean country, does not have a comprehensive reference national olive collection. A report published 30 years ago indicated the presence of four Lebanese varieties, but the confusion regarding these varieties was discussed then and is still prevalent today hindering prospects for conserving and exploring the national germplasm. This study assessed the existing in situ diversity of old Lebanese olive groves using simple sequence repeat (SSR) and amplified fragment length polymorphisms (AFLP). Trees (140) were collected from 14 groves located in the four traditional olive areas. Analysis was based on 22 polymorphic alleles produced from six SSR primers, and on 135 polymorphic AFLP fragments. SSR and AFLP did not yield consistent results in terms of level of polymorphism, with SSR detecting higher variation than AFLP. On the other hand, both clustered trees based on geographic provenance. AFLP coefficient of similarity between trees ranged between 0.70 and 0.99 indicating the possible presence of varieties since some values fall within reported intervarietal ranges of 0.65 to 0.84. SSR unveiled the presence of nine different genotypes: four corresponding each to a provenance and five consisting of single trees characterized by the presence of less frequent alleles with an average of 0.33. Results also revealed a high incidence of clones greater than 90% in three of the four provenances. The findings of this study point for the need to plan for a sampling strategy that takes into consideration geographic provenances.     

Molecular evolution and diversity in Bacillus anthracis as detected by amplified fragment length polymorphism markers.

Bacillus anthracis causes anthrax and represents one of the most molecularly monomorphic bacteria known. We have used AFLP (amplified fragment length polymorphism) DNA markers to analyze 78 B. anthracis isolates and six related Bacillus species for molecular variation. AFLP markers are extremely sensitive to even small sequence variation, using PCR and high-resolution electrophoresis to examine restriction fragments. Using this approach, we examined ca. 6.3% of the Bacillus genome for length mutations and ca. 0.36% for point mutations. Extensive variation was observed among taxa, and both cladistic and phenetic analyses were used to construct a phylogeny of B. anthracis and its closest relatives. This genome-wide analysis of 357 AFLP characters (polymorphic fragments) indicates that B. cereus and B. thuringiensis are the closest taxa to B. anthracis, with B. mycoides slightly more distant. B. subtilis, B. polymyxa, and B. stearothermophilus shared few AFLP markers with B. anthracis and were used as outgroups to root the analysis. In contrast to the variation among taxa, only rare AFLP marker variation was observed within B. anthracis, which may be the most genetically uniform bacterial species known. However, AFLP markers did establish the presence or absence of the pXO1 and pXO2 plasmids and detected 31 polymorphic chromosomal regions among the 79 B. anthracis isolates. Cluster analysis identified two very distinct genetic lineages among the B. anthracis isolates. The level of variation and its geographic distribution are consistent with a historically recent African origin for this pathogenic organism. Based on AFLP marker similarity, the ongoing anthrax epidemic in Canada and the northern United States is due to a single strain introduction that has remained stable over at least 30 years and a 1,000-mile distribution.     

分子标记在啤酒花种质资源研究中的应用

该论文利用分子生物学中常用的DNA分子标记对世界各地现存的野生和栽培的啤酒花种质资源遗传多样性研究的应用进展做一综述。通过查阅和研读20世纪90年代以来发表的各类文献进行归纳总结。发现DNA分子标记相比形态学标记和细胞学标记具有结果准确、稳定的特点,常用的分子标记技术有RAPD、RFLP、ISSR、SSR、AFLP、EST等;研究发现北美洲的啤酒花遗传多样性要高于欧洲的啤酒花,基因变异程度也相对较高;野生啤酒花的基因序列具有丰富的基因多样性,可在分子杂交遗传育种中作为一个种质去改善栽培品种的某些不良性状。因此,利用分子标记研究啤酒花的遗传多样性将对啤酒花的优良育种提供理论指导和技术支持,目前较为理想的技术是SSR和AFLP。     

Assessment of genetic variation within Indian mustard (Brassica juncea) germplasm using random amplified polymorphic DNA markers

Genetic diversity among 45 Indian mustard (Brassica Juncea L.) genotypes comprising 37 germplasm collections, five advance breeding lines and three improved cultivars was investigated at the DNA level using the random amplified polymorphic DNA (RAPD) technique. Fifteen primers used generated a total of 92 RAPD fragments, of which 81 (88%) were polymorphic. Of these, 13 were unique to accession 'Pak85559'. Each primer produced four to nine amplified products with an average of 6.13 bands per primer. Based on pairwise comparisons of RAPD amplification products, Nei and Li's similarity coefficients were calculated to evaluate the relationships among the accessions. Pairwise similarity indices were higher among the oilseed accessions and cultivars showing narrow ranges of 0.77-0.99. An unweighted pair-group method with arithmetic averages cluster analysis based on these genetic similarities placed most of the collections and oilseed cultivars close to each other, showing a low level of polymorphism between the accessions used. However, the clusters formed by oilseed collections and cultivars were comparatively distinct from that of advanced breeding lines. Genetically, all of the accessions were classified into a few major groups and a number of individual accessions. Advanced breeding lines were relatively divergent from the rest of the accessions and formed independent clusters. Clustering of the accessions did not show any pattern of association between the RAPD markers and the collection sites. A low level of genetic variability of oilseed mustard was attributed to the selection for similar traits and horticultural uses. Perhaps close parentage of these accessions further contributed towards their little diversity. The study demonstrated that RAPD is a simple and fast technique to compare the genetic relationship and pattern of variation among the gene pool of this crop.