Amplified-fragment Polymorphism Research Articles

Water temperature and chlorophyll a density drive the genetic and epigenetic variation of Vallisneria natans across a subtropical freshwater lake.

Plant genetic diversity differs in habitat's oscillations, especially species distributed under heterogeneous environmental conditions. Freshwater ecosystems are vulnerable to biotic and abiotic impacts, which affect the genetic and epigenetic variations in aquatic plants. The extent of environmental heterogeneous attributes can be examined based on genetic and epigenetic variations. Such variations under environmental gradient can provide evidence for understanding the correlations between rapid environmental changes and species evolution. In this study, we performed amplified fragment polymorphism length and methylated-sensitive amplified polymorphism analysis to depict the genetic and epigenetic variations of Vallisneria natans in a subtropical lake. Results showed that this species maintained a relatively high genetic diversity (mean H E = 0.320, I = 0.474, PPL = 85.93%) and epigenetic variation (mean eH E = 0.282, eI = 0.428, ePPL = 83.91%). Water body temperature and chlorophyll a density were positively correlated to the genetic and epigenetic variations. The clonal generates of V. natans depicted a relative high methylation level and shew ancestral scenario between the genet and the second clonal generation. These findings revealed that species diversity is unevenly distributed under environmental heterogeneity, even at a fine geographic scale. Environmental characteristics in relation to temperature and chlorophyll a should be considered in the analysis of the genetic and epigenetic variations. Additionally, epigenetic variations between genets and ramets should be considered with caution when applied to analysis of other aquatic species.

Exploring genetic diversity and disease response of cultivated rice accessions (Oryza spp.) against Pyricularia oryzae 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 and relatedness of the mangrove Rhizophora mangle L. (Rhizophoraceae) using amplified fragment polymorphism (AFLP) among locations in Florida, USA and the Caribbean

Rhizophora mangle L. is a widespread mangrove species in the Western Hemisphere. Mangrove habitat loss and their importance to coastal and reef ecosystems make greater understanding of their genetic structure useful for conservation and management. An amplified fragment polymorphism (AFLP) analysis was performed on samples from Florida and the Caribbean to discover the genetic structure present. R. mangle had variable genetic diversity not related to latitude; P ranged 7 %–92 %. Some other factor, perhaps human impact, has caused low genetic diversity in some populations. Across Florida R. mangle populations varied in genetic diversity with less diversity (Gst = 0.195) and greater gene flow on the Atlantic coast (Nm =2.07) than on the Gulf coast (Gst = 0.717, Nm = 0.197). Gene flow between Caribbean islands was low (Nm = 0.386) compared to continental populations (Nm = 1.40), indicating that long distance dispersal is not common between islands. Analysis of molecular variance (AMOVA) analysis showed significant deviations from Hardy-Weinberg expectations at the level of region among subpopulations and overall genetic difference among subpopulations for R. mangle. One implication for management is that small continental populations and island populations may be genetically isolated and distinct from each other.

Toxigenic profile and AFLP variability of Alternaria alternata and Alternaria infectoria occurring on wheat

The objectives of this study were to evaluate the ability to produce alternariol (AOH), alternariol monomethyl ether (AME) and tenuazonic acid (TA) by A. alternata and A. infectoria strains recovered from wheat kernels obtained from one of the main production area in Argentina; to confirm using AFLPs molecular markers the identify of the isolates up to species level, and to evaluate the intra and inter-specific genetic diversity of these two Alternaria species. Among all the Alternaria strains tested (254), 84% of them were able to produce mycotoxins. The most frequent profile of toxin production found was the co-production of AOH and AME in both species tested. TA was only produced by strains of A. alternata. Amplified fragment polymorphism (AFLPs) analysis was applied to a set of 89 isolates of Alternaria spp (40 were A. infectoria and 49 were A. alternata) in order to confirm the morphological identification. The results showed that AFLPs are powerful diagnostic tool for differentiating between A. alternata and A. infectoria. Indeed, in the current study the outgroup strains, A. tenuissima was consistently classified. Characteristic polymorphic bands separated these two species regardless of the primer combination used. Related to intraspecific variability, A. alternata and A. infectoria isolates evaluated seemed to form and homogeneous group with a high degree of similarity among the isolates within each species. However, there was more scoreable polymorphism within A. alternata than within A. infectoria isolates. There was a concordance between morphological identification and separation up to species level using molecular markers. Clear polymorphism both within and between species showed that AFLP can be used to asses genetic variation in A. alternata and A. infectoria. The most important finding of the present study was the report on AOH and AME production by A. infectoria strains isolated from wheat kernels in Argentina on a semisynthetic media for the first time. Also, specific bands for A. alternata and A. infectoria have been identified; these may be useful for the design of specific PCR primers in order to differentiate these species and to detect them in cereals.

Worldwide genotyping of castor bean germplasm (Ricinus communis L.) using AFLPs and SSRs

Worldwide genetic diversity in 200 individuals comprising 41 castor bean accessions was assessed using amplified fragment polymorphisms (AFLPs) and simple sequence repeats (SSRs). We found that, despite surveying five continents and 35 countries, genetic diversity in castor bean germplasm is relatively low (overall He = 0.126 for AFLPs and 0.188 for SSRs) compared to estimates of genetic diversity in other plant species. Our data also show no geographic structuring of genotypes across continents or countries within continents. An assessment of the congruence between AFLP and SSRs indicates a low correlation (R2 = 0.19) between the two data sets, but each marker class nonetheless shows similar patterns of low-genetic diversity and a lack of geographic structure. Our data do suggest that SSRs yield a higher percentage of polymorphic loci, higher heterozyosity and a greater range of genetic distances, and are therefore more informative than are AFLPs on a locus-by-locus basis. Based on comparisons with numerous other plant species, we suggest that the lower genetic variation in this worldwide collection may be due to one or more factors including: sampling strategies that have not captured the full extent of genetic variation in the species; artifactual variation due to long-term germplasm storage and seed regeneration; or intense selection followed by domestic cultivation of a limited number of castor bean genotypes, which are widely propagated for their horticultural and agro-economic value.

Alteration of methylation profiles in distinct cell lineages of the layers during vegetative propagation in carnations (Dianthus caryophyllus)

In vegetatively propagated plants, branches of periclinal chimera give rise to bud mutants, or “sports”, which have been used to breed new cultivars. Here, we have examined DNA methylation profiles in the cells of the L1 and L2+3 layers from single plants of vegetatively propagated carnations. The band patterns of methylation-sensitive amplified fragment polymorphism (M-AFLP) of the genomic DNAs in the L1 and L2+3 layers of the carnation cultivar, “White Sim”, vegetatively propagated over the past 50 years, were completely different. The bud mutant, “White Mind”, obtained from “White Sim” about 25 years ago, also had very different M-AFLP patterns to the parental “White Sim” line. The cultivar, “Red”, which was separated from “Satisfaction” as a bud mutant two years ago, showed similar patterns to “Satisfaction” in each corresponding layer. However, we were able to detect minor, but significantly different M-AFLP patterns between the cultivars, “Red” and “Satisfaction”. The number of bands that differed between these cultivars was larger in the L2+3 layers than in the L1 layers. The results indicate that the DNA methylation profiles differ between each cell layer derived from distinct cell lineages of vegetatively propagated plants, and that changes in these profiles occur frequently and accumulate in the cells of the L2+3 layers, rather than in the L1 layer.

Phylogenetic analysis of Bacillus thuringiensis based on PCR amplified fragment polymorphisms of flagellin genes.

Phylogenetic analysis of 35 strains of Bacillus thuringiensis was performed based on PCR amplified fragment polymorphisms of flagellin genes (PCR-AFPF). From coefficient matrix, the genetic distance coefficient D among them ranged from 0.14286 to 0.81818. Ten clusters were divided by the taxonomic threshold of 0.42 from the UPGMA dendrogram, demonstrating that the genetic dissimilarities existed among 35 strains of B. thuringiensis corresponding to 35 different serovars. This method could provide a fast, convenient and accurate way to classify all subspecies of B. thuringiensis including strains that can not be classified using the H-antigen method.

Bacillus cereus cellulitis from contaminated heroin

Concern exists over recent unexplained deaths among intravenous drug users. This report describes a patient with crepitant cellulitis who was admitted complaining of severe pain in the right forearm. Ultrasonography demonstrated gas in the tissues and he was referred for early surgical debridement of the arm. He was treated with intravenous benzyl penicillin, gentamicin and metronidazole and made a full recovery. Aspirate samples grew Bacillus cereus, morphologically similar to the isolate obtained from a sample of the patient's own heroin. Antibiogram and API 50CHB profiles were also similar. Further typing included 'H' flagellar serotyping, which found both blood and heroin strains to be non-typable, and amplified fragment polymorphism analysis, which showed that the strains were indistinguishable. Genotyping of two selected genes from B. cereus confirmed almost certain identity between the two strains. This case illustrates the potential virulence of B. cereus when inoculated into tissues, and to our knowledge, is the first report to demonstrate a conclusive microbiological link between contaminated heroin and serious sepsis in a drug user due to B. cereus.

Groat yield of naked and covered oat

Near-isogenic covered (CN 18941) and naked (CN 18942) lines of the oat (Avena sativa L.) cultivar NO 141-1 were developed to determine if the contrasting spikelet morphologies of the two lines affected groat yields differentially. Molecular markers verified the high level of genetic similarity between the two lines. Their groat yields were not significantly different under field conditions. Key words: Hulless oats, Hulled oats, Avena sativa L., near- isogenic lines, groat yield, amplified fragment polymorphism

Genetic diversity and its relationship to hybrid performance in maize as revealed by RFLP and AFLP markers

The challenge to maize breeders is to identify inbred lines that produce highly heterotic hybrids. In the present study we surveyed genetic divergence among 13 inbred lines of maize using DNA markers and assessed the relationship between genetic distance and hybrid performance in a diallel set of crosses between them. The parental lines were assayed for DNA polymorphism using 135 restriction fragment length polymorphisms (RFLPs) and 209 amplified-fragment polymorphisms (AFLPs). Considerable variation among inbreds was detected with RFLP and AFLP markers. Moreover AFLPs detect polymorphisms more efficiently in comparison to RFLPs, due to the larger number of loci assayed in a single PCR reaction. Genetic distances (GDs), calculated from RFLP and AFLP data, were greater among lines belonging to different heterotic groups compared to those calculated from lines of the same heterotic group. Cluster analysis based on GDs revealed associations among lines which agree with expectations based on pedigree information. The GD values of the 78 F1 crosses were partioned into general (GGD) and specific (SGD) components. Correlations of GD with F1 performance for grain yield were positive but too small to be of predictive value. The correlations of SGDs, particularly those based on AFLP data, with specific combining-ability effects for yield may have a practical utility in predicting hybrid performance.