Biotechnology Research and Innovation最新文献

Ependymoma, a rare pediatric tumor originating from ependymal cells located in the lining of ventricular surfaces in the brain, presents great challenges in treatment despite advances in neurosurgical techniques. In order to identify new molecular biomarkers that could improve clinical management and outcomes, we have used RNA-seq to profile the Whole Transcriptome of three ependymoma samples and one normal control brain tissue, producing a total of 2.5 Gigabases of sequencing data. Different protein-coding gene databases were interrogated for known annotated genes to calculate RPKM and Fold Changes (FGs) for each clustered gene. Using this approach, we were able to identify Differentially Expressed Genes (DEGs) in ependymomas. KEGG pathways and Gene Ontology (GO) categories enriched in the DEGs were analyzed using Enrichr, and protein interaction networks were then built using MetaCore™. Thirty differentially expressed protein-coding genes associated with neurogenesis were identified by TaqMan Real-Time PCR, 17 of these showing statistically significant differential expression. Based on these results, we have identified IGF-2 as highly over-expressed in ependymomas and that this is due to loss of DNA methylation in its promoter region. In conclusion, we believe that some of these genes, specially IGF-2, may be of clinical importance, opening new avenues for disease management and new therapies.

The present study was performed to assess the effects of maleic acid (MA), ethylenediaminetetraacetic acid (EDTA) amendments on biochemical parameters of 7 days old seedlings of Hordeum vulgare L. under Co(II) stress. Application of MA, EDTA, Co(II) alone significantly reduced the root length and shoot length, whereas combination of MA and Co(II), and EDTA and Co(II) enhances the root and shoot length. The antioxidative enzymes such as superoxide dismutase, catalase, guaiacol peroxidase, ascorbate peroxidase and glutathione reductase were significantly enhanced under application of MA, EDTA, Co(II) alone, while their combinations reduced the activities of all the antioxidative enzymes. Multivariate regression analysis and β-regression coefficients showed that β₁(Co) and β₂(MA) have positive regression on the antioxdative enzymes, while β₃(Co × MA) have negative regression with the antioxidative enzymes. Similarly, β₁(Co) and β₂(EDTA), and β₃(Co × EDTA) showed the similar trend as that of Co and MA. Malondialdehyde (MDA) content is increased under Co stress, whereas MA and EDTA alone or in combination with Co reduce the MDA content. Multiple linear regression analysis also showed that MDA content is significantly enhanced under β₁(Co) stress, whereas in combination with MA or EDTA decreases the MDA content.

Tuberculosis (TB) is among the top ten causes of mortality worldwide and has high prevalence in developing countries. The dissemination of efficient and low-cost diagnosis tools able to identify its latent form, e.g. the delayed hypersensitivity reaction, is of great importance to accomplish the target of TB eradication. Recent studies have shown the potential of specific Mycobacterium tuberculosis immunodominant antigens, CFP10 and ESAT6, as substitutes of tuberculin skin test. The purpose of this study was to optimize the recovery and purification of recombinant CFP10 and ESAT6 from Escherichia coli inclusion bodies using central composite design. The production of CFP10 and ESAT6 in bioreactor presented yields of 233 and 121 mg L⁻¹, respectively, after extraction under optimized conditions: biomass concentration of 15 g per 150 mL of sonication buffer, using 12 cycles of disruption and 7 cycles of solubilization, followed by affinity chromatography purification and removal of endotoxins by the micellar method. The identity of the antigens was confirmed by mass spectrometry and their immunoreactivity after recovery and purification was confirmed by Western blot. These results demonstrate the suitability of the preparation methods in the development of a TB diagnosis kit with potential to be diffused in high TB burden countries.

Setaria viridis, a C₄ monocot, was proposed as a model plant for studies on the response to stress conditions. Water-deficit ranks among the top three most devastating stresses and its importance will likely increase in the scenario of climate change. The aim of this work was to evaluate physiological and molecular water-deficit responses of S. viridis subjected to different conditions. Principal component analysis highlighted the physiological differences between vegetative and reproductive stages of S. viridis, as well as the differences between two methods of water-deficit induction: polyethylene glycol and air-drying. Network interactions were observed in distinct developmental stages and water-deficit induction methods tested, allowing classification of root and shoot fresh weight and non-photochemical quenching as the best physiological parameters to group the networks. Variations in the gene expression patterns of delta 1-pyrroline-5-carboxylate synthase 2 (SvP5CS2), Dehydrin 1 (SvDHN1) and the transcription factors WRKY DNA-binding domain 1 (SvWRKY1), dehydration-responsive element-binding protein 1 class C (SvDREB1C) and NAC protein 6 (SvNAC6) were observed. Among these genes, it was observed two expression patterns predominant during water-deficit: inducible (SvDHN1 and SvNAC6) and repressed (SvP5CS2, SvWRKY1 and SvDREB1C) genes. SvDHN1 showed the highest expression level in all the conditions tested. PEG treatment during the reproductive stage promoted the upregulation of the five marker genes in roots. The discriminative analysis suggested that the physiological and molecular responses in S. viridis adjusted according to the evaluated water-deficit conditions, especially after PEG treatments, indicating that the PEG method of water-deficit may better replicate field conditions.

This research explored the lignocellulolytic potential of native strains (Hipoxylon sp, Curvularia kusanoi, Trichoderma sp and Aspergillus fumigatus) isolated from natural substrates. Their sequences were deposited in the GenBank, and it‘s lignocellulosic potentialities were determined quantitatively from kinetics of cellulase and ligninase production in wheat straw, and grass hay. All evaluated strains showed high cellulolytic potential and laccase activity where only found in the C. kusanoi L7 strain. The new strains where co-cultivated with a reference strains of Trichoderma viride to determinate the capacity of joint growth, to know if they can be used together for a better degradation of the cell wall. A three-phase purification system was used to isolate the C. kusanoi laccase where a preparation 10 times purer than the initial crude was obtained, with an enzymatic activity of 2800 U/L, a specific activity of 544.74 U/g and a yield higher than 100%, which showed a more stable behavior at alkaline pH than at acid pH. The purify enzymes showed an optimum range of activity between 30 and 40 °C, and a high thermal stability. It is concluded that the isolated strains have a high degradation capacity and the C. kusanoi L7 strain stands out as the one with the greatest potential when expressing in addition to the cellulolytic complex the enzymes involved in the degradation of lignin.

A novel technology was developed to obtain a high amount of pyroligneous acid (PA) rich in organics. PA was obtained by atmospheric pyrolysis of a mixture of pine, spruce and fir wood particles, analyzed by gas chromatography-mass spectrometry (GC–MS) and evaluated for in vitro antibacterial and antifungal activity. Several microbial inhibitory compounds were observed in PA. Antimicrobial activity of PA was studied at both acidic pH (3.7) and neutral pH (7.0) of the liquid. Neutralized PA showed higher antibacterial activity than acidic PA against 5 pathogenic bacterial strains, and Minimum Inhibitory Concentration (MIC) obtained with neutralized PA were 0.3125% (v/v), 0.3125% (v/v), 0.625% (v/v), 0.3125% (v/v), 0.3125% (v/v) for Escherichia coli, Enterobacter aerogenes, Pseudomonas aeruginosa, Listeria monocytogenes and Enterococcus faecalis, respectively. For fungal strains, acidic PA was found to be more effective than neutralized PA. The highest activity was against Trametes versicolor, followed by Aspergillus niger and Aspergillus fumigatus. The MIC of acidic PA with which fungal inhibition was seen was 0.125% (v/v) for T. versicolor and A. fumigatus, and 0.25% (v/v) for A. niger. Hence, the novel technology was found to be effective to produce a high yield of PA (40–45 wt% of dry biomass), rich in antimicrobial compounds, and the PA is proposed as a potential alternative to antibiotics, preservatives and/or chemical disinfectants that are currently in use.

Brazilian agricultural biotechnology has seen great advances in recent decades, especially in the development of GM crops, including soybean, cotton, and maize, which has placed Brazil in second place since 2013 in the ranking of countries with the greatest GM-cultivated area. However, patenting these technologies is somewhat more restrictive in Brazil than in other countries, such as the USA and Japan, especially concerning isolated biological material from nature. Hence, the intellectual protection of crops in Brazil is encompassed by sui generis rights and/or the patenting of only the development process. Given the current scenario and the importance of biotechnology for the Brazilian agriculture sector, it is necessary to deeply study the patent system for recently developed technologies to identify opportunities for enterprises and national institutes to act in this area. The application of novel biotechnological strategies to agriculture will contribute to the expanding agriculture sector and become part of the solution to global challenges. Through this study, we can identify the major companies developing and protecting their agrobiotechnologies. Additionally, a more detailed analysis verifies that although there are some restrictions in Brazilian laws, GM patent applicants find ways to obtain intellectual protection for the tools they use in the development of GM crops, which include regulatory sequences, gene constructs and production methodologies. Mechanisms to stimulate investment in Brazilian research companies and public policies must be consolidated, allowing investment and public–private partnerships in this sector, with the aim of applying biotechnological knowledge and turn it into products demanded by society.

Patent protection has been chosen as a strategy to protect new developments in molecular biology such as novel genes and proteins. A way to ensure the protection of genetic inventions is to claim a set of sequences that are associated with the described genetic sequences in terms of structure and/or biological activity, in a genus claim. Clearly, achieving an effective patent protection for proteins and genetic sequences is a real challenge for an Intellectual Property manager, considering the unpredictability of biological sciences and the diversity in current patent law and patent office guidance in each territory. This paper seeks to study the Brazilian patent office procedures about genus claims for biological sequences while comparing them with two other national/regional offices. To achieve this result, we initially present the concepts, followed by the current requirements and the barriers to obtain genus claims for biological sequences in the legal framework and patent office prosecution of Brazil, the European Union, and the United States. Subsequently, we study the impacts of these regulations in the scope of claim protection in each territory. This is done by comparing patent documents with the same priority granted in each of these offices in order to analyze the extension of the owner's rights for biological sequences. Understanding the logic that supports the examination procedures in the three studied offices will be important to subsidize the legal protection for gene-based inventions. Therefore, this would support the development of a patent system that can provide satisfactory safeguard for the results of investments in biotechnology Research and Development initiatives.

In a search of new-fangled antitumor chemotherapeutic agent, the studies are elucidating the enzymes, contributing to the metabolic reprogramming and their potential as therapeutic targets. This study was an endeavor to explore the potential of enzyme pterin deaminase from cyanide degraders. In preliminary and secondary screening, potent cyanide degraders were selected and estimated for pterin deaminase activity. The intracellular crude extract containing enzyme activity was found to be higher in potential strain Bacillus cereus AM12. The optimization conditions for the production of pterin deaminase from this selected strain was studied with the help of Box–Behnken design. The determination coefficient of the model (0.91) was highly significant and indicative of goodness of fit. This design found to be ideal for the selected parameters, such as temperature of 35 °C, pH 8, 0.005 M of pterin and 20% of glucose which yielded maximum production of pterin deaminase (9.99 U/ml). This study might make a major breakthrough in the field of medicine as well as in pharmaceutical compositions when the biological significance of this distinct enzyme is proven.