Endophytic bacteria are an important part of different functions in plants that lead to plants’ production characteristics as well as their stress response mechanisms. Endophytic bacterial diversity was analyzed in this study to describe 16S rRNA variability and changes in the leaves of drought-tolerant and drought-susceptible wheat when growth under in vitro conditions. A metagenomic analysis was applied and a pilot exploratory study was performed to prove this type of analysis as applicable to tracking endophytic bacterial diversity changes when a drought stress is applied to an in vitro culture of wheat. The study showed that the changes in the bacterial endophytes’ variabilities associated preferentially with the drought stress varietal characteristics of the analyzed wheat instead of the applied stress conditions.
Bacterial contamination of semen is an important factor connected to the health status of bulls that may significantly affect semen quality for artificial insemination. Moreover, some important bovine diseases may be transmitted through semen. Up to now, only a very limited number of complex studies describing the semen microbiome of bulls have been published, as many bacteria are hard to cultivate using traditional techniques. The 16S rRNA high-throughput sequencing strategy allows for the reliable identification of bacterial profiles of bovine semen together with the detection of noncultivable bacterial species. Fresh samples from Holstein Friesian breeding bulls (n = 55) were examined for the natural variability in the present bacteria. Semen doses were selected randomly from Slovak Biological Services in Nitra, Slovak Republic. The most predominant phyla within the whole dataset were Firmicutes (31%), Proteobacteria (22%), Fusobacteria (18%), Actinobacteria (13%) and Bacteroidetes (12%). Samples of semen were divided into two separate clusters according to their microbiome compositions using a cording partition around a medoids analysis. Microbiomes of the first cluster (CL1) of samples (n = 20) were based on Actinobacteria (CL1 average = 25%; CL = 28%) and Firmicutes (CL1 = 38%; CL2 = 27%), while the second cluster (CL2; n = 35) contained samples characterized by a high prevalence of Fusobacteria (CL1 = 4%; CL2 = 26%). Some important indicator microbial groups were differentially distributed between the clusters.
Background and objectives: A metagenomic approach was applied to analyze endophytic bacterial diversity changes in Amaranthus cruentus L. subjected to mutagenesis-based breeding. Findings: High-throughput sequencing approach of 16S rRNA variability analysis was used for monitoring of the changes of endophytic bacterial diversity in selected M generations and varieties Ficha and Pribina of Amaranthus cruentus L.A total of 15,683,430 clusters were generated in the sequencing and 15,085,381 of them passed filter. Average cluster density was 646 clusters per mm 2 . A 95%, 97% of sequences were Q >30. All sequences were classified into the 44 orders that were identified from all the analyzed genotypes of Amaranthus. Metagenomic libraries of Ficha and Pribina showed the similar profiles of identified bacterial orders and that are different from the profiles of M1 and M7 generations after gamma radiance treatment. Conclusions: Endophytic bacteria diversity was higher in the starting variety and final new variety of Amaranthus when comparing to the M generations. Based on the laboratory conditions of the growth of analyzed plant material, it supposes that endophytic bacterial abundance differences are a result of temporary changes in the amount of bacterial endophytes and the metagenomic limitation of their identification. Significance and novelty: Metagenomic analysis provides a sensitive approach that facilitates the study of the endophytic bacteria changes in the leaves of plant after gamma radiation mutagenesis. K E Y W O R D S Amaranthus cruentus L., gamma radiance mutants, metagenomic analysis *The authors contributed to the study equally
Several types of allergies are currently known and are characterized by an exaggerated response of the immune system to substances from various sources called allergens. One of them is a food allergy, which is becoming more common in the population. For this reason, it is necessary to describe the issue from several aspects including genomic variability of plant allergens. The objective of this study was to analyse intraspecific variability of Bet v 1 of 10 different varieties of apple species (Malus domestica Borkh.). BBAP technique for genomic determination of the presence of Bet v 1 homologs at the DNA level was performed. Degenerate primers that anneal a variable and conserved part of PR-10 protein homologues genes were used in the analyse. Amplicons were generated and formed relatively monomorphic profiles, indicating the stability of the given isoforms of Bet v 1 within the selected apple varieties. To evaluate the potential allergenicity of selected varieties further studies on another molecular level such as a comparison of gene expression of the PR-10 family members and their protein expression levels are needed.
Different types of allergies became a part of life of many people around the world. The research activities connecting to allergens are actually not oriented only for protein and immunological interactions, but to the genomic and transcriptomic background of them, too. Analysis and description of genomic variability of allergens in plant food resources will help to manage the allergen based strategies in the future. Here, the bioinformatic approach was used to develop and validate the specific primers for genomic screening of polymorphism of profilins (Profilin Based Amplicon Polymorphism; PBAP) and vicilins (Vicilin Based Amplicon Polymorphism; VBAP) among the legumes. The alignment of existing public databases data for these allergens in the group of legumes was performed. Subsequently, specific primers were designed and their ability to generate polymorphic amplicons were tested for three legumes – bean, lentil and chickpeas. In all cases, amplicons were generated and polymorphism was detected in all three species for profilin as well as for vicilin.
The aim of this study was clarifying the relation between genetic diversity of edible honeysuckle (Lonicera kamtschatica) and the major group of biologically active substances as total polyphenols content (TPC) including antioxidant activity (AO). Fruits of edible honeysuckle becomes more and more popular, especially in Europe. The current status of research on polyphenolic compounds in the berries of edible honeysuckle and their biological effects, including recommended utilization, are reviewed.The biological material including 14 cultivars of the edible honeysuckle (´Zoluška´, ´Amfora´, ´Pruhonický 44´, ´Vasilijevsky´, ´Moskovskaja´, ´Vojtek´, ´Sinoglaska´, ´Altaj´, ´Lipnická´, ´Kamčadalka´, ´Sinaja Ptica´, ´Fialka´, ´Modrý Triumf´, and ´Leningradský velikán´) originated from Czech republic (Žabcice near Brno). The content of TPC and AO were determined by location and its soil-climatic conditions and these environmental circumstances determines the RAPD profiles of analysed honeysuckle acessions, too. DPPH method was used to analyze AO and Folin-Ciocalteu method was used to determine TPC. The results of experiment showed that the highest value of AO was determined at the cultivars ´Zoluška´ (81.04 mg.L-1) and the lowest was measured in ´Kamčadalka´ (54.122 mg.L-1). On the contrary, the highest content of TPC was determined at the cultivar ´Kamčadalka´ (51.09 mg.L-1) and the lowest value was measured at the cultivar ´Pruhonický 44´ (21.65 mg.L-1). Phylogenetic trees were similar in genetic distance. The content of TPC and AO were not statistically significant in relation to cultivar. The analyzed cultivars of the edible honeysuckle were separated in 4 clusters according to used primers. In both gel images, the amplicon size ranged from 100 to 1,500 bp. We found that genetic diversity was partially related to content of total polyphenolic substances and antioxidant activity. Based on phylogenetic trees we have stated that variety ´Lipnická´, ´Sinoglaska´, ´Altaj´, ´Leningradský velikán´, ´Modrý Triumf´, ´Sinaja Ptica´ and ´Kamčadalka´ were grouped in the similar cluster. The highest genetic distance was determined at the variety ´Vasilijevskaja´ and ´Amfora´. In the same way, there were variety ´Vojtek´, ´Fialka´ and ´Zoluška´.
A collection of Amaranthus cruentus accessions was analysed to compare the characteristics of different dinucleotide core sequences of microsatellites repeats. ISSR analysis was performed to obtain corresponding genetic fingerprints and based on them, to construct dendrograms for every individual primer used in the study. A total of eighteen accesions were analysed. Dinucleotide sequence of (CA)6 repetition anchored by GG bases resulted in a total of 65 DNA fragments and 100-percent polymorphism. This marker has differ completely only 10 from all of the analysed genotypes. Dinucleotide sequence of (CT)8 repetition anchored by AC bases has resulted in the amplification of totally 97 fragments and 84.62 % of polymorphism. Only seven accesions were different among themselves when regarding this microsatellite core sequence. Dinucleotide sequence of (GA)6 repetition anchored by CC bases has resulted in the amplification of totally 171 DNA fragments and 100 % polymorphism. Not all the genotypes were separated based on this marker, two of them - Ames 5638 RRC 1139 and Ames 5648 RRC 1148 - possess the same profile of amplified ISSR fragments. The most distant genotype separated by all of the used primers was a landrace PI 511876 Huatle originated in Mexico.
Rosa canina, L. is widely used for medicinal purposes as well as in food industry where it is a valuable source, bioactive compounds and natural colorants. Actually, no specific method is recommended as suitable one for DNA extraction from rose hips. The aim of the study was to compare three commercial and three non-commercial methods to extract total genomic DNA from rose hips hyphanthium. Four methods are based on the precipitation in principle and two methods are based on resin-binding. Extracted DNA was proved for the effectivity in following PCR. In total, six different DNA isolations was performed for differently heat processes rose hips -fresh hyphanthium, 2-weeks frozen hyphanthium, dried hyphanthium (50 °C) and boiled hyphanthium (100 °C). The amplification parameters of 500 bp chloroplast gene amplicon were evaluated. Obtained amounts of extracted DNA was very variable not only for every individual method used but for individual treatment of samples, too. In general, non-commercial method provided higher amount of extracted DNA, but the A260/280 ratio was lower. When regarding the processing treatment of the samples, high differences were found among the samples untreated by heat and those that were dried or boiled for three of the used extraction methods. All the samples were positive for amplification, but different amounts of amplified product were obtained. The comparison of data for concentrations of extracted DNA and concentrations of amplified product showed large differences when regarding the achieved purity of DNA in extraction.
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