The aim of this study was to characterize extracts from the leaves of Ginkgo biloba L. from selected Slovakian localities in terms of the content of bioactive constituents, antioxidants and their antimicrobial properties. The results indicated that the content of antioxidants was sample-specific, and this specificity was statistically significant. Ginkgo biloba L. from the locality of Košice had the best activity determined by the free radical scavenging activity (DPPH) (1.545 mg Trolox equivalent antioxidant capacity (TEAC)/g fresh matter (FM)) as well as the molybdenum-reducing antioxidant power (35.485 mg TEAC/g FM) methods. The highest content of total polyphenols (2.803 mg gallic acid equivalent (GAE)/g FM) and flavonoids (4.649 μg quercetin equivalent (QE)/g FM) was also detected in this sample. All samples of G. biloba leaf extracts showed significant antimicrobial activity against one or more of the examined bacterial species, and Staphylococcus aureus subsp. aureus CCM 2461 was found to be the most susceptible (minimal inhibition concentration MIC50 and MIC90 values of 64.2 and 72.2 µg/mL, respectively). Based on the results it was concluded that Ginkgo biloba L. extracts can be used as antimicrobial and antioxidant additives. Selected miRNA-based molecular markers were used to examine the environmental adaptability of Ginkgo biloba L. An almost-complete genotype clustering pattern based on locality was determined in the analysis that involved a species-specific gb-miR5261 marker. Morphologically specific exemplar, cv. Ohatsuki, was excluded.
MicroRNAs (miRNAs) are a class of non-coding RNAs about 20-24 nucleotides long. They play an important role in the gene regulation at the post-transcriptional level. They affect the plant genome response to environmental stress. The miRNA-based molecular markers is type of functional markers reported in very few plants. However, the information connected to the evaluation of genotypes by this type of markers within a single species is missing. Considering the stability, polymorphism, functionality and transferability potential of miRNA-based markers, the research was conducted to apply selected types of them (miR156b, miR408a and the combined type of miR156b/miR408a) for the genotyping analysis of eight flax genotypes of different origin together with the morphology analyses. A total of 145 miRNA loci were identified, of which 19 were unique. The highest numbers of miRNA loci (57) and unique fragments (9) as well as the highest percentage of polymorphism and the extent of polymerase chain reaction (PCR) amplification of miRNA fragments have been observed with the combination of miR156b-F and miR408-F markers. By means of the miRNA markers has been recorded the unique profile of the miRNA loci for individual accessions. The morphology study has shown that the genotypes are the same in the expression of selected morphological traits despite the different use and different places of origin. However, we have identified an interface between some of morphological traits and miRNA-based markers for genotyping the genetic resources of flax. By mutually linking these two types of markers, we were able to determine unique genotypes of flax.
Two types of molecular markers based on randomly amplified DNA by RAPD-based assay and amplified microRNA by miRNA-based assay were applied for the genotyping of five accessions of Silybum marianum (L.) Gaertn. It was also verified the effectiveness of the isolation of genomic DNA by three commercially available isolation kits and two similar complex methods and their applicability for this medicinal herb. None of the commercial isolation kits provided the genomic DNA of sufficient quality and quantity. Applied originally designed CTAB method and modified CTAB method for purpose of DNA isolation from medicinal plants allowed to isolate DNA of required quality and of sufficient yield. RAPD-based DNA fingerprints, allowed to distinguish the individual genotypes of Silybum marianum. MicroRNA-based markers showed the cross-genera transferability potential and displayed sufficient level of polymorphism. Both types of molecular markers could be used as suitable tool for genotyping of milk thistle. However, because of the size of miRNA amplicons is the efficiency of miRNA-based markers PCR amplifications preferred and consequently less DNA quality-dependent which may be advantageous in view of the content of undesirable secondary metabolites in medicinal plants.
MicroRNAs represent small non-coding RNAs that play important role in regulating gene expression under various biotic and abiotic stresses as well as in different developmental stages of plants. They are involved in wide variety of biological and metabolic pathways of plants. The research was focused on the potential of selected miRNA-based molecular markers (miR156 and miR168) to map the polymorphism level of flax genome in the selected developmental stages (flower bud -floweringboll development) as a reflection of the activity of specific miRNAs in these flax organs and tissues. The miRNA polymorphism was evaluated on 8 flax genotypes by miRNA-based molecular marker assays and data were supported by the morphology measurements on buds, flowers, petals and developing bolls. Extent of PCR amplification of miRNA fragments ranged from 40 bp to 200 bp (miR168a-F/miR-R primer pair) or 35 bp to 120 bp (miR156b-F/miR-R primer pair), respectively. MiRNAbased primers amplified in total 196, respectively 158 miRNA loci. Based on the results can be concluded that the representation of miR168 loci have increased according to developmental stages ascending (stage of bud, flower and boll). In overall miRNA156b loci profile in all three analyzed developmental stages is possible to observe the increase of miRNA loci amplification almost in all genotypes. Taking into account the genetic background of genotypes, based on the peak analysis profile of amplified miRNA loci, implemented by GeneTools software, was possible to identified unique loci in linseed and intermediate genotypes in individual developmental stages. Have been found out that only the genotype Bolley Golden differ from other genotypes in both analysis (molecular and morphological) if the
MicroRNAs (miRNAs) are a class of 19 - 24 nucleotide long non-coding RNAs derived from hairpin precursors, regulating various biological, metabolic and developmental processes at the post-transcriptional level. Many of the known miRNAs are evolutionary conserved across diverse plant species and function in the regulatory control of fundamentally important biological processes. It is known that exogenous plant miRNAs specifically target approximately 30% of protein-coding genes in mammals. The research was focused to analyze the occurrence of selected families of miRNAs (miR156, miR168 and miR171) in less used species but nutritionally important plant food resources (flax and medlar) and medicinal plant (milk thistle). The analyses were done by two individual approaches, by (a) miRNA-based molecular markers - as a novel type of functional markers and (b) qualitative Real-Time PCR. The expression pattern of selected miRNAs was analyzed depending on various plant tissues and developmental stages. Results have confirmed the significance and reliability of novel type of markers based on miRNA molecules as well as the species-specific and tissues-specific expression patterns of plants miRNAs. Significant polymorphism profile of miR156b was detected in various flax tissues of genotypes varying in the content of alpha-linolenic acid. Conclusions indicate that the variable behavior of the miRNA molecules, depending on various factors, may reflect the variability of the gene expression regulation of the human genome. The exploitation of the background of miRNA functioning within different species and plant tissues will help us to understand the molecular machinery as well as the regulatory mechanisms involved in the expression of miRNAs in plants and consequently in human genome.
Flax (Linum usitatissimum L.) is one of the oldest domesticated plants — it was cultivated as early as in ancient Egypt and Samaria 10,000 years ago to serve as a source of fiber and oil, whence it later spread around the world. Compared with other plants, the flax genome consists of a high number of repetitive sequences, middle repetitive sequences and small repetitive sequences of nucleotides. The aim of the study was to analyze the stability of the existing trinucleotides motifs of microsatellite DNA of the flax genome (genotype Kyivskyi), growing in the Chernobyl conditions. The Chernobyl area is the most extensive “natural” laboratory suitable for the study of radiation effects. Over the last 20 years, the researches collected important knowledge about the effects of low and high radiation doses on the DNA isolated from the plant material growing on the remediated fields near Chernobyl and the plant material from fields contaminated by radioactive cesium 137Cs and strontium 90Sr. Using eight pairs of microsatellite primers, we successfully amplified the samples from the remediated fields. For each primer in the control samples and remediated samples, we detected 1 to 3 fragments per locus, each in size up to 120 to 250 base pairs. The applied microsatellite primers confirmed the monomorphic condition of microsatellite loci.
The most suitable method of total DNA extraction still remains the crucial step for many plant species, although there are many different protocols and commercial kits for DNA isolation. In this study, five different extraction protocols were analysed to find out the most appropriate method for DNA extraction from Hedera helix L. This species has numerous medical and pharmaceutical uses and is also characterized by antioxidant effects on human body. In spite of its wide medical utilization, it belongs to those plant species, where the genomic information is very limited. Comparing of different protocols resulted in the yield of extracted DNA that has ranged from 6.3 to 487 ng µl -1. The purity of extracted DNA has ranged from 1.4 up to 2.0 A260/A280. All the extraction methods used in this study were evaluated not only in term of quantity and purity of DNA but also its functionality in the restriction endonuclease digestion and polymerase chain reaction based downstream analysis was performed.
In rational nutrition, vegetables play an important role due to their high biological and low energy value. The most widespread vegetables in our country belong to root vegetables. They are grown mainly for bulbs, corms, rhizomes, fleshy roots, and hypocotyl tubers. Root vegetables can be eaten raw or cooked. Carrot (Daucus carota L.) is a basic representative of root vegetables. For the most valuable components counts beta-carotene – the major component of total carotenoids. This paper evaluates changes in total carotenoids, refractometric dry matter, and gravimetric dry matter in three varieties of carrot (Kamaran F1, Komarno F1, Romosa) grown in soil and climatic conditions ex-situ in Nitra. We have evaluated roots grown in non-fertilized soil, soil after application of manure, horticultural compost, and their combinations. The results show that the variants fertilized with compost and a mixture of compost and manure had the most considerable influence on the synthesis and content of total carotenoids as well as the content of dry matter and refractometric dry matter
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