Water shortage is a major environmental stress that causes the generation of reactive oxygen species (ROS). The increase in ROS production induces molecular responses, which are key factors in determining the level of plant tolerance to stresses, including drought. The aim of this study was to determine the expression levels of genes encoding MAPKs ( MAPK3 and MAPK6 ), antioxidant enzymes ( CAT , APX and GPX ) and enzymes involved in proline biosynthesis ( P5CS and P5CR ) in Triticum aestivum L. seedlings in response to short-term drought conditions. A series of wheat intervarietal substitution lines (ISCSLs) obtained by the substitution of single chromosomes from a drought-sensitive cultivar into the genetic background of a drought-tolerant cultivar was used. This source material allowed the chromosomal localization of the genetic elements involved in the response to the analyzed stress factor (drought). The results indicated that the initial plant response to drought stress resulted notably in changes in the expression of MAPK6 and CAT and both the P5CS and P5CR genes. Our results showed that the substitution of chromosomes 3B, 5A, 7B and 7D had the greatest impact on the expression level of all tested genes, which indicates that they contain genetic elements that have a significant function in controlling tolerance to water deficits in the wheat genome.

Background: Quantitative PCR (qPCR) is one of the most common and accurate methods of gene expression analysis. However, the biggest challenge for this kind of examinations is normalization of the results, which requires the application of dependable internal controls. The selection of appropriate reference genes (RGs) is one of the most crucial points in qPCR data analysis and for correct assessment of gene expression. Because of the fact that many reports indicate that the expression profiles of typically used RGs can be unstable in certain experimental conditions, species or tissues, reference genes with stable expression levels should be selected individually for each experiment. In this study, we analysed a set of ten candidate RGs for wheat seedlings under short-term drought stress. Our tests included five 'traditional' RGs (GAPDH, ACT, UBI, TUB, and TEF1) and five novel genes developed by the RefGenes tool from the Genevestigator database. Results: Expression stability was assessed using five different algorithms: geNorm, NormFinder, BestKeeper, Ref-Finder and the delta Ct method. In the final ranking, we identified three genes: CJ705892, ACT, and UBI, as the best candidates for housekeeping genes. However, our data indicated a slight variation between the different algorithms that were used. We revealed that the novel gene CJ705892, obtained by means of in silico analysis, showed the most stable expression in the experimental tissue and condition. Conclusions: Our results support the statement, that novel genes selected for certain experimental conditions have a more stable level of expression in comparison to routinely applied RGs, like genes encoding actin, tubulin or GAPDH. Selected CJ705892 gene can be used as a housekeeping gene in the expression analysis in wheat seedlings under short-term drought. The results of our study will be useful for subsequent analyses of gene expression in wheat tissues subjected to drought.

Enterobacter aerogenes LU2 was isolated from cow rumen and recognized as a potential succinic acid producer in our previous study. Here, we present the first complete genome sequence of this new, wild strain and report its basic genetic features from a biotechnological perspective. The MinION singlemolecule nanopore sequencer supported by the Illumina MiSeq platform yielded a circular 5,062,651 bp chromosome with a GC content of 55% that lacked plasmids. A total of 4,986 genes, including 4,741 protein-coding genes, 22 rRNA-, 86 tRNA-, and 10 ncRNA-encoding genes and 127 pseudogenes, were predicted. The genome features of the studied strain and other Enterobacteriaceae strains were compared. Functional studies on the genome content, metabolic pathways, growth, and carbon transport and utilization were performed. The genomic analysis indicates that succinic acid can be produced by the LU2 strain through the reductive branch of the tricarboxylic acid cycle (TCA) and the glyoxylate pathway. Antibiotic resistance genes were determined, and the potential for bacteriocin production was verified. Furthermore, one intact prophage region of length ~31,9 kb, 47 genomic islands (GIs) and many insertion sequences (ISs) as well as tandem repeats (TRs) were identified. No clustered regularly interspaced short palindromic repeats (CRISPRs) were found. Finally, comparative genome analysis with well-known succinic acid producers was conducted. The genome sequence illustrates that the LU2 strain has several desirable traits, which confirm its potential to be a highly efficient platform for the production of bulk chemicals. Enterobacter aerogenes LU2 is a gram-negative, wild bacterium that was isolated from cow rumen as a part of environmental screening for succinic acid-producing bacteria identification. Succinic acid (SA) plays an important role as a metabolic intermediate in the rumen by increasing propionate production, a crucial energy source for the ruminant 1,2. Anaerobic conditions and the presence of carbon dioxide, methane and trace amounts of hydrogen create an excellent environment for the biosynthesis of succinate by some bacteria existing in the rumen 3. In reports of the U.S. Department of Energy (DOE) from 2004 and 2010, SA was recognized as one of the top 10 most promising C4-building chemical platforms for the production of high-value commodity and specialty chemicals with great industrial potential 2,4. Succinate is widely applied as an additive in food, pharmaceuticals, detergents, solvents, and surfactants as well as in biodegradable polymer production 5,6. Until recently, industrial production of succinate was based on chemical synthesis. However, petroleum-based production of SA from n-butane through maleic anhydrate requires the use of high pressure, high temperature and costly catalysts 7. Therefore, because of the current global trend regarding sustainable development, including the support of green technologies, rational waste biomass management and pollution-reducing standards, the bio-based production of...

Background: Succinic acid (SA), a valuable chemical compound with a broad range of industrial uses, has become a subject of global interest in recent years. The bio-based production of SA by highly efficient microbial producers from renewable feedstock is significantly important, regarding the current trend of sustainable development. Results: In this study, a novel bacterial strain, LU2, was isolated from cow rumen and recognized as an efficient producer of SA from lactose. Proteomic and genetic identifications as well as phylogenetic analysis were performed, and strain LU2 was classified as an Enterobacter aerogenes species. The optimal conditions for SA production were 100 g/L lactose, 10 g/L yeast extract, and 20% inoculum at pH 7.0 and 34 °C. Under these conditions, approximately 51.35 g/L SA with a yield of 53% was produced when batch fermentation was conducted in a 3-L stirred bioreactor. When lactose was replaced with whey permeate, the highest SA concentration of 57.7 g/L was achieved with a yield and total productivity of 62% and 0.34 g/(L*h), respectively. The highest productivity of 0.67 g/(L*h) was observed from 48 to 72 h of batch fermentation, when E. aerogenes LU2 produced 16.23 g/L SA. Conclusions: This study shows that the newly isolated strain E. aerogenes LU2 has great potential as a new biocatalyst for producing SA from whey permeate.

Fusarium populations were investigated on 53 samples of wheat grains and husks collected approximately three weeks before harvest in 53 wheat fields in southeastern Poland. A limited area of sample collection was chosen intentionally to avoid the effect of climate and weather variability. The study was conducted to assess the occurrence of 13 Fusarium species using species-specific PCR assays separately on grains and husks of winter wheat. The obtained data suggest that husks could take a protective role of wheat grain against Fusarium spp. The incidence of Fusarium species decreased in grains vs. husks from 29 to 100%. While Fusarium species were present in husks at 11.32% and less, they were absent in the grain. The presence of Fusarium species on husks is inversely proportional to the percentage reduction of Fusarium spp. in grain. There was a correlation of the presence of certain species of Fusarium in husks and in grains. The number of Fusarium species found on husks was about three times higher in comparison to wheat grain. In conclusion, the presented data indicate the importance of Fusarium populations analysis on wheat husk in seeds pathological studies.

Non-coding RNAs (ncRNAs) have been considered as unimportant additions to the transcriptome. Yet, in light of numerous studies, it has become clear that ncRNAs play important roles in development, health and disease. Long-ignored, long non-coding RNAs (lncRNAs), ncRNAs made of more than 200 nucleotides have gained attention due to their involvement as drivers or suppressors of a myriad of tumours. The detailed understanding of some of their functions, structures and interactomes has been the result of interdisciplinary efforts, as in many cases, new methods need to be created or adapted to characterise these molecules. Unlike most reviews on lncRNAs, we summarize the achievements on lncRNA studies by taking into consideration the approaches for identification of lncRNA functions, interactomes, and structural arrangements. We also provide information about the recent data on the involvement of lncRNAs in diseases and present applications of these molecules, especially in medicine.

Background: Quantitative PCR (qPCR) is one of the most common and accurate methods of gene expression analysis. However, the biggest challenge for this kind of examinations is normalization of the results, which requires the application of dependable internal controls. The selection of appropriate reference genes (RGs) is one of the most crucial points in qPCR data analysis and for correct assessment of gene expression. Because of the fact that many reports indicate that the expression profiles of typically used RGs can be unstable in certain experimental conditions, species or tissues, reference genes with stable expression levels should be selected individually for each experiment. In this study, we analysed a set of ten candidate RGs for wheat seedlings under short-term drought stress. Our tests included five ‘traditional’ RGs (GAPDH, ACT, UBI, TUB, and TEF1) and five novel genes developed by the RefGenes tool from the Genevestigator database.Results: Expression stability was assessed using five different algorithms: geNorm, NormFinder, BestKeeper, RefFinder and the delta Ct method. In the final ranking, we identified three genes: CJ705892, ACT, and UBI, as the best candidates for housekeeping genes. However, our data indicated a slight variation between the different algorithms that were used. We revealed that the novel gene CJ705892, obtained by means of in silico analysis, showed the most stable expression in the experimental tissue and condition. Conclusions: Our results support the statement, that novel genes selected for certain experimental conditions have a more stable level of expression in comparison to routinely applied RGs, like genes encoding actin, tubulin or GAPDH. Selected CJ705892 gene can be used as a housekeeping gene in the expression analysis in wheat seedlings under short-term drought. The results of our study will be useful for subsequent analyses of gene expression in wheat tissues subjected to drought.

In recent years, zebrafish (ZF) has been increasingly applied as a model in human disease studies, with a particular focus on cancer. A number of advantages make it an attractive alternative for mice widely used so far. Due to the many advantages of zebrafish, modifications can be based on different mechanisms and the induction of human disease can take different forms depending on the research goal. Genetic manipulation, tumor transplantation, or injection of the pathogen are only a few examples of using ZF as a model. Most of the studies are conducted in order to understand the disease mechanism, monitor disease progression, test new or alternative therapies, and select the best treatment. The transplantation of cancer cells derived from patients enables the development of personalized medicine. To better mimic a patient’s body environment, immune-deficient models (SCID) have been developed. A lower immune response is mostly generated by genetic manipulation but also by irradiation or dexamethasone treatment. For many studies, using SCID provides a better chance to avoid cancer cell rejection. In this review, we describe the main directions of using ZF in research, explain why and how zebrafish can be used as a model, what kind of limitations will be met and how to overcome them. We collected recent achievements in this field, indicating promising perspectives for the future.