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Linum album is an herbaceous and medicinal plant that has important lignan such as podophyllotoxin (PTOX). PTOX has antiviral and anticancer properties. Since the chemical synthesis of PTOX is an expensive process, production of PTOX using cell and cultures of linum species is a cost-effective alternative approach. Various strategies have been employed to increase the production of secondary metabolites in cell cultures. In this study, we have verified the effect of chitosan on cell growth, PTOX production in 1, 2, 3 and 5 days after treatment. Cells elicited with chitosan for 5 days yielded the highest amount of PTOX. To study mechanism of chitosan action, expression of phenylalanine ammonio-lyase (PAL), cinnamoyl-CoA reductase (CCR), cinnamyl alcohol dehydrogenase (CAD) and pinoresinol lariciresinol reductase (PLR) genes were investigated. The expression of genes were increased, reaching a peak at 3 day after treatment. Chitosan up-regulate the production of PTOX, by effecting on gene expression of PTOX biosynthesis pathway.

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Sheath blight, caused by Rhizoctonia solani AG1-IA, is one of the most destructive disease. Conventional methods of disease control using fungicides may develop new problems. Therefore, understanding molecular mechanisms of plant–pathogen interaction is necessary to adopt effective approaches for managing the disease. Here for the first time, by using bioinformatics tools and RT-PCR analysis and sequencing confirmed the presence of a Magnaporthe oryzae Avr-pita gene orthologous sequence designated as Rhiz-pita1 gene in three different geographic isolates of R. solani AG1- IA( A2,R1 and T2) genome. SignalP program predicted a secretion signal upstream of Rhiz-pita1 gene. Nucleotide sequences of 5' region of Rhiz-pita1 gene from geographical isolates showed 99% identity in exons and 100% in introns which are characteristics of fast evolving effector proteins. Also, 98% homology between Rhiz-pita and M.oryza-pita1gene suggests that Rhiz-pita encodes an effector protein. Howevere, more researchs are necessary to confirm of this suggestion. Keywords: Rhizoctonia solani, signal peptide Rice blast , Effector

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Abstract: ATP sulfurylase (ATPS) is widely distributed in all living organisms. Several different physiological roles have been proposed for ATPS in different species, including sulfate assimilation, sulfate reduction and pyrophosphate recycling. Also, ATP sulfurylase has many different industrial and laboratory applications. The aim of this study was to clone and express the gene that producing the recombinant ATPS protein from an Iranian strain of Geobacillus. After Isolation and identification of Geobacillus kaustophilus strain, DNA genomic was extracted. ATPS gene was amplified from genomic DNA by using a couple of specific primers for interested gene. PCR product of ATPS gene was observed as an 1188bp band on agarose gel. Then the PCR product was purified and cloned into the cloning vector. The ATPS band was sequenced after cloning and result of homology search in the NCBI database confirmed that the cloned gene was ATPS. The ATPS gene was subcloned in expression pET28a plasmid. Expression of recombinant ATPS protein in E. Coli BL21 (DE3) was analyzed using SDS-PAGE gel. Analysis of expressed ATPS protein on SDS-PAGE gel revealed a band at 47.5 KD. Using ATP luminescence method for measuring enzymatic activity of the protein showed that the recombinant protein is active. This is the first study on cloning, expression and enzymatic activity of the ATPS gene from the Geobacillus kaustophilus bacteria.

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Aims: Most parts of the pastoralist's livelihood depend on income from livestock in the rangelands, but drought in rangelands has negative impacts on the ecosystem, cycle of nomadic pastoralists, livestock production, and their family's living. Recognizing the signs of drought effects in the nomadic environment can play a significant role in rangeland management. The present study was conducted to identify and investigate drought signs in Gonbad-e Kavous County, Golestan Province, Iran.
Materials & Methods: The study's statistical population consisted of 183 nomadic pastoralists in 10 customary systems, 100 of them were sampled and classified by random method. The data gathering tool was a researcher-made questionnaire. Drought signs were investigated in three dimensions: vegetation, soil, livestock, and herd. In order to ensure the validity of the questionnaire, the views of the experts of the Natural Resources Bureau and Cronbach's alpha method were used for more reliability with an average of 0.717.
Findings: The results showed that nomadic pastoralists assessed three factors forage quality degradation, increasing bare land, and livestock production as the most important indicators of drought effects in pastureland. Findings indicate a positive and significant relationship between the pastoralist's viewpoints about the signs of drought effect in the pasture with the variable of the people who depend on them. The results showed a significant difference between respondents' views on gender and education level in the context of drought effects.

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Objective: The ITPA gene is responsible to remove free deaminated purine nucleotides of ITP, dITP and XTP from nucleotide pool of the cells. It seems that dysfunction in its activity, not only can increas the base substitution mutations frequency but also can works as a contrived factor to creating instability in genetic materials of the cells. There are several reports about the existence of structural and numerical genetic instability in the K562 cell genome. In this research, we examined the expression of ITPA gene as a possible contrived factor in observed genetic instability of this cell line. Materials and Methods: To evaluate the expression of target gene semi-quantitative RT-PCR technique was used. Then to examine the functionality of gene products, its cDNAs were cloned and their sequences were determined. Their proteins products were predicted using available bioinformatics soft wares and the results were compared. Results: The result of structural prediction of second mRNA showed that it has ability to encode a protein which has inability in substrate binding and also in its normal enzymatic activity. With regard to the fact that enzymatic activity of protein is dependent on the dimer formation, the function of hetero-dimer enzyme is changed. Therefore the catalytic activity of ITPase is predicted to be abnormal and it can be considered as a contrived factor for creating genetic instability in K562 cell line. Conclusion: The study of gene expression showed that ITPA gene is expressed in moderate level compared to GAPDH expression as an internal control in K562 cell. Two types of transcripts were detected in this line. One of them was the normal product of splicing process of primary hnRNA, but the second one contained a 51 nucleotides deletion in the mRNA coding region. It seems, this transcript is the product of a rare splicing process in this line.

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Objective: E1A oncoprotein of adenovirus type 5 is a regulatory factor which controls transcription of other adenovirus genes. This protein promotes both viral genom replication and host cell transformation by altering the function of certain important cellular proteins such as p21 and Rb. The aim of the present study was to constantly reduce the expression of E1A gene in HEK 293 cell line by RNAi technique in order to analyse the effects of this suppression on these cells. Materials and Methods: The U6 promoter and shRNA regions from the control and E1A specific siRNA coding plasmid as pSP-81 and pSP81-E1A were subcloned into pcDNA3.1. Then these constructs were transfered into the HEK 293 cancerous cells using lipofection method and successfully transfected cell colonies were selected based on neomycin antibiotic resistance. Changes in E1A gene expression were analysed by RT-PCR technique after selection process. Results: Final analysis showed no obvious difference in E1A gene expression level in the suppresed and control groups, upon transfection with the constructed plasmids. In order to examine the possible influence of cloning procedure on the function of U6 promoter, cells were transfected with Dr. Hacker’s original plasmids, but no inhibition of E1A gene expression was observed again. The results of sequencing revealed existence of a mutation in the siRNA target region for the E1A gene sequence. Conclusion: These results illustrated that no considerable suppression has been occurred by repeating Dr. Hacker’s expriment, even with application of very effective lipofection method. To examine the sequence of the E1A gene, the PCR product of the 13s region of the gene was sequenced. Sequencing revealed existence of a point mutaion in the siRNA target region. It seems that observed impaired interference could be attributed to this mutation in the E1A gene of the studied cells.

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In recent years, many studies have been performed on for use in various of science. The proper design and synthesis of these has a direct impact on their -chemical properties and their applications, especially in the field of biological sciences. There are several methods for magnetic synthesis. One of the simplest and most efficient methods for synthesis of magnetic is a chemical co-precipitation method, but of magnetic is one of the limitations of this method. In this study, various protocols for the synthesis of magnetic by co-precipitation method and silica coating of magnetic were performed and the effect of different factors such as the type of alkaline compound, the use of , temperature and in dispersion, aggregation of magnetic nanoparticles and their stability in aqueous solutions was investigated. Finally, a simple and reproducible protocol for magnetic synthesis with appropriate size distribution and high dispersion in aqueous solutions was optimized for use in biological applications.

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The unique physicochemical properties of nanoscale plasmonic materials have attracted considerable attention in the fabrication of hybrid nano-bio structures because of their promising applications in biosensing, imaging, and controlled-release drug delivery. The purpose of this study was the synthesis of functionalized gold nanorods (GNRs) to both reduce the toxicity and increase the biocompatibility for further applications such as the design of a therapeutic nanocarrier for nucleic acid delivery to cancerous cells. In this study, GNRs were prepared by seed-mediated method and their surface was modified by polystyrene sulfonate (PSS) polymer. Then, peptide-functionalized GNRs was fabricated via ligand exchange method through the Au-S bond. The CTAB-GNRs and functionalized nanostructures were characterized using ultraviolet-visible spectrophotometry, transmission electron microscopy (TEM), and zeta potential measurement. Finally, the cytotoxicity effects of functionalized GNRs on Hela cells were studied by MTT assay. The optimal concentration of PSS and peptide, which did not cause any aggregation and morphological perturbations of the nanostructure were obtained 50μM and 1mM respectively. The survival percentage of treated Hela cells significantly increased by surface modification of GNRs with PSS and functionalization with peptide compared to CTAB-GNRs. While LC50 of functionalized GNRs was calculated 50nM, treated cells with the same concentrations of CTABGNRs survived less than 20%. Functionalization of GNRs increases its biocompatibility and improves applications of this nanostructure as a therapeutic carrier in cancerous cells.

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Objective: One of the most significant damages to the genetic material is oxidative deamination of DNA and free nucleotides in the cell pool. Incorporation of deaminated purine nucleotides such as inosine triphosphate (ITP, dITP) into the DNA can increase the frequency of base substitution mutation. it has been suggested that presence and accumulation of these rough nucleotides can lead to genetic instability which is the perquisite of different types of diseases or cancers. Inosine triphosphate pyrophosphates (ITPase) encoded by ITPA gene, is responsible for protecting the cells by omitting deaminated purines from the free nucleotide pool. The objective of this study was to examine the possible dysfunction of ITPA gene activity as an important factor in genetic background predisposing to chromosomal disorders and malignancies such as CML. Materials and Methods: ITPA gene expression study performed on 23 CML patients and 21 controls using semi-quantitative RT-PCR technique and the expression level of GAPDH gene was used as an internal control. Unusual variants obtained from cDNA amplification of ITPA gene were cloned. Results: Our results showed a significant reduction of ITPA gene expression in CML patients in comparison to the controls. Two types of transcripts were produced in addition of expected transcript in some samples. One of them had a 123 nucleotide and the other had 77 nucleotide deletions in their open reading frame. Conclusion: Due to decreased expression of ITPA gene; it seems that the function of ITPase is not normal in CML patients. Therefore the altered expression of this gene can be considered as an additive factor for genetic instability in these patients.

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Abstract: Hyperglycemia is a major cause of diabetes. Hyperglycemia-induced endothelial dysfunction is generally believed to be the basis of diabetic vascular complications such as retinopathy, nephropathy and cardiovascular diseases. The most important molecules in endothelial cells that can sense elevated level of glucose and transmit signals into the cell are G protein-coupled receptors (GPCRs).
In the present study, according to bioinformatics analysis of genomic sequences between healthy and patient individuals, two G proteins GPR182 and CALCRL were selected and their expression level were examined in hyperglycemic and normal conditions in HUVEC as a model of vascular endothelial cells at different glucose concentrations and various time intervals. In addition, the effects of hyperglycemia on cell viability and cell cytotoxicity were assessed by MTT and LDH assay respectively and also morphological changes by immunohistochemistry.
Overall our data reveal a probable role for GPR182 and CALCRL in hyperglycemia-induced endothelial dysfunction. Thus, they could be developed as a potential molecular targets for the endothelial dysfunction therapy.

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Atherosclerosis is a chronic vascular disease and remains the leading cause of death and morbidity worldwide. Endothelial dysfunction is an important factor in the progression of atherosclerotic disease. Increased expression of cell adhesion index genes and decreased cell-binding proteins lead to abnormal endothelial function. These molecular changes are one of the most important indicators of endothelial cell dysfunction and the progression of atherosclerosis. CXCR3 is a G-protein-coupled chemokine receptor expressed by endothelial cells. The role of the receptor CXCR3 and its ligands in endothelial cells and heart disease is not yet fully understood. In this study, we evaluated the effect of CXCR3 downregulation on the expression level of adhesion (I-CAM-1, V-CAM-1), tight junction (TJP1), related to endothelial dysfunction.
In order to reduce the expression of the CXCR3 gene, the RNA-cleaving DNAzyme was used against the mRNA of the CXCR3 gene. DNAzyme was transfused into HUVEC cells by TurboFect^TM. After confirmation of decreased CXCR3 gene expression, RNA extraction and cDNA synthesis were performed and then the expression of markers was evaluated by RT-qPCR technique.
Our result was showed the expression level of I-CAM-1 and V-CAM-1 were showed significant up-regulation in transfected cells compared with control cells, while the TJP1 gene was not showed significant change. It seems that reducing the CXCR3 gene expression could induce endothelial dysfunction through the change of adhesion markers genes expression. Therefore, this receptor can be considered as a potential molecular target for a better understanding of the mechanism of atherosclerosis.

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Aim: In this study, the antioxidant properties of hydrolyzed protein from longtail tuna dark muscle with commercial enzymes (alcalase, alkaline protease, and evatase) were investigated.
Materials & Methods: Protein hydrolysates from tuna dark muscle were prepared by different enzymes Degree of hydrolysis (DH) was performed by TCA technique. The five aliquots at 60, 180, 240, 300, and 360 min were gathered during hydrolysis. The antioxidant activity of aliquots was monitored by in vitro assays (DPPH inhibition ability and Ferric (Fe3+) reducing power).
Findings: The antioxidant activities of protein hydrolysate from tuna dark muscle (TDM) increase with increasing time and DH. Alcalase hydrolyzed protein (AHP) generally showed higher antioxidative activity than evatase hydrolyzed protein (EHP) and alkaline protease hydrolyzed protein (APHP). Among the samples (concentration 3 mg.ml^-1), AHP at 360 min significantly exhibited the highest ability to scavenge DPPH radical (72.6 %). Furthermore, AHP and APHP significantly showed a minimum IC50 value of 1.1 mg.ml^-1 at 240 and 360 min hydrolysis. APHP significantly exhibited the highest ferric reducing power of 0.83 at 300 min and 0.76 at 240 min. AHP and APHP significantly showed the highest ferric reducing power of 0.74 at 360 min (p < 0.05).
Conclusion: This study confirmed that protein hydrolysate from TDM could be a good source of antioxidant peptides. In addition, the antioxidant activity of hydrolyzed protein relay on protease type and hydrolysis condition.
 

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Transforming growth factor beta (TGF-β), is a small homodimeric signaling protein. The TGF-β isoforms (TGFβ1, β2 and β3) are involved in many cellular processes including growth inhibition, extracellular matrix remodeling, tissue development, cell migration, invasion and immune regulation. For research aims, TGFβs are overexpressed using recombinant eukaryotic cell or bacterial expression systems. For achieving an efficient purification of TGF-β by immobilized metal ion affinity chromatography (IMAC), a histidine tag was placed either at the C-terminal (C-TGFβ) or N-terminal (N-TGFβ) region of the sequence and the effect of His-tag on TGF-β structure has been studied by computational tools. Proteins 3D structures were modeled using MODELLER software and molecular dynamics simulation of native TGF-β and modelled proteins, N-TGFβ and C-TGFβ were studied in water by GROMACS package. Protein dynamics modeling indicated that the His-tag attached at the C-terminus but not at the N-terminus of the TGF-β can affect the fluctuations of amino acids and protein structure. It is concluded that the C-terminal tagging may cause distortion and misfolding in the structure.

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 Multiple sclerosis (MS) is one of the most common autoimmune diseases in Iran and the world. To date, many drugs have been developed to control the progression of MS as a chronic inflammatory disease of the central nervous system. Rituximab is a chimeric mouse-human monoclonal antibody that binds to the CD20 receptor on the surface of B cells and induces apoptosis. Today, Numerous studies have confirmed the increasing role of non-coding RNAs in regulating the expression of genes and molecular processes, including apoptosis. Furthermore, bioinformatic analysis results indicate that TUG1 LncRNA is differentially expressed in MS patients. Thus, In the present study the possible role of TUG1 in regulating rituximab mechanism of action and apoptosis induction was experimentally investigated. To do this, specific DNAzyme against TUG1 was designed and transfected into Raji cells in the presence or absence of the drug. After transfection, RNA extraction and cDNA synthesis were performed. Then, the expression of target genes was examined by Real-Time PCR technique. The results showed an increase in CD20 expression and a decrease in SMAD2 expression levels. Furthermore, decreased TUG1 gene expression led to an increase in apoptosis and cell accumulation in the G1 phase. It seems that TUG1 expression level can play a significant role in CD20 expression in B cells and therefore on the therapeutic efficacy of rituximab.
 

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Objective: Evaluating the effects of p-benzoquinone and hydroquinone on the RUNX2 expression and osteoblastic differentiation of human marrow derived mesenchymal stem cells (MSCs). Materials and Methods: Bone marrow MSCs obtained by cultivating marrow mononuclear cells, were exposed to 10μM of either p-benzoquinone or hydroquinone. Following chemical treatment, RUNX2 gene expression was assessed by Real-time RT PCR 1, 6, 24 and 48 hours later and osteogenic differentiation was analyzed using alizarin red and alkaline phosphatase staining methods on days 7 and 14 after ostegenic induction. Results: RUNX2 expression was significantly elevated (up to approximately 8 times) due to chemical exposure but the applied chemicals exert no considerable effect on MSCs osteogenic differentiation. Conclusion: According to the literature, despite the necessity of RUNX2 overexpression on the induction of osteogenic differentiation, but it is not sufficient for osteogenesis to occure so increase in RUNX2 expression observed in our study is not the indicator of the induced osteogenic differentiation. Instead, this elevated expression could be the sign of increased activity of the canonical Wnt signaling pathway thereby its involvement in the development of AML due to exposure to benzene and its metabolites. Moreover, this augmented expression of RUNX2 in MSCs can indicate the RUNX2 overexpression in myeloid progenitors as an expected similar effect of exposure to benzene and its metabolites to contribute in myeloid malignancies developed due to benzene exposure.