Search results for: biosynthetic gene clusters

Authors: Ruchi Yadav, Shivani Pandey, Prachi Srivastava

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Microarrays provide a rich source of data on the molecular working of cells. Each microarray reports on the abundance of tens of thousands of mRNAs. Virtually every human disease is being studied using microarrays with the hope of finding the molecular mechanisms of disease. Bioinformatics analysis plays an important part of processing the information embedded in large-scale expression profiling studies and for laying the foundation for biological interpretation. A basic, yet challenging task in the analysis of microarray gene expression data is the identification of changes in gene expression that are associated with particular biological conditions. Careful statistical design and analysis are essential to improve the efficiency and reliability of microarray experiments throughout the data acquisition and analysis process. One of the most popular platforms for microarray analysis is Bioconductor, an open source and open development software project based on the R programming language. This paper describes specific procedures for conducting quality assessment, visualization and preprocessing of Affymetrix Gene Chip and also details the different bioconductor packages used to analyze affymetrix microarray data and describe the analysis and outcome of each plots.

Keywords: microarray analysis, R language, affymetrix visualization, bioconductor

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Authors: Ahmad Ali Shahid, Mukhtar Ahmed

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The demand for long staple fiber having better strength and length is increasing with the introduction of modern spinning and weaving industry in Pakistan. Work on gene discovery from developing cotton fibers has helped to identify dozens of genes that take part in cotton fiber development and several genes have been characterized for their role in fiber development. Sucrose synthase (SuS) is a key enzyme in the metabolism of sucrose in a plant cell, in cotton fiber it catalyzes a reversible reaction, but preferentially converts sucrose and UDP into fructose and UDP-glucose. UDP-glucose (UDPG) is a nucleotide sugar act as a donor for glucose residue in many glycosylation reactions and is essential for the cytosolic formation of sucrose and involved in the synthesis of cell wall cellulose. The study was focused on successful Agrobacterium-mediated stable transformation of SuS gene in pCAMBIA 1301 into cotton under a CaMV35S promoter. Integration and expression of the gene were confirmed by PCR, GUS assay, and real-time PCR. Young leaves of SuS overexpressing lines showed increased total soluble sugars and plant biomass as compared to non-transgenic control plants. Cellulose contents from fiber were significantly increased. SEM analysis revealed that fibers from transgenic cotton were highly spiral and fiber twist number increased per unit length when compared with control. Morphological data from field plants showed that transgenic plants performed better in field conditions. Incorporation of genes related to cotton fiber length and quality can provide new avenues for fiber improvement. The utilization of this technology would provide an efficient import substitution and sustained production of long-staple fiber in Pakistan to fulfill the industrial requirements.

Keywords: agrobacterium-mediated transformation, cotton fiber, sucrose synthase gene, staple length

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Authors: Xiaohan Bookman, Xiaoyan Zhu

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The outputs of non-linear feed-forward neural network are positive, which could be treated as probability when they are normalized to one. If we take Entropy-Based Principle into consideration, the outputs for each sample could be represented as the distribution of this sample for different clusters. Entropy-Based Principle is the principle with which we could estimate the unknown distribution under some limited conditions. As this paper defines two processes in Feed-Forward Neural Network, our limited condition is the abstracted features of samples which are worked out in the abstraction process. And the final outputs are the probability distribution for different clusters in the clustering process. As Entropy-Based Principle is considered into the feed-forward neural network, a clustering method is born. We have conducted some experiments on six open UCI data sets, comparing with a few baselines and applied purity as the measurement. The results illustrate that our method outperforms all the other baselines that are most popular clustering methods.

Keywords: feed-forward neural network, clustering, max-entropy principle, probabilistic models

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Authors: Neda Menbari, Ramin Mehdiabadi

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Background: breast cancer remains a critical global health issue, constituting a leading cause of cancer-related mortality in women. MicroRNAs (miRs) are natural RNA molecules that play an important role in cellular processes and regulate post-transcriptional gene expression. MiR-216b-5p is a miR that acts as a tumor suppressor. The expression levels of FoxM1 and miR-216b-5p in malignant and control cells have been evaluated by quantitative polymerase chain reaction (qPCR) technique and flow cytometry. Results: the results of this study revealed a significant downregulation of miR-216b-5p in cancerous cells compared to the control MCF-10A cells (P=0.0004). Interestingly, the expression of miR-216b-5p exhibited an inverse relationship with key clinical indicators such as tumor size, grade, and lymph node invasion. Conclusion: The study's findings showed the prognostic value of miR-216b-5p levels in breast cancer, and its reduced expression correlates with unfavorable tumor characteristics. This research recommends performing more studies on the role of FoxM1 and miR-216b-5p in breast cancer pathology which potentially paving the way for targeted therapeutic interventions.

Keywords: breast cancer, gene expression, FOXM1, microRNA

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Authors: Samira B. R. Prado, Paulo R. Melfi, Beatriz T. Minguzzi, João P. Fabi

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Fruit softening is the main change that occurs during papaya (Carica papaya L.) ripening. It is characterized by the depolymerization of cell wall polysaccharides, especially the pectic fractions, which causes cell wall disassembling. However, it is uncertain how the modification of the two main pectin polysaccharides fractions (water-soluble – WSF, and oxalate-soluble fractions - OSF) accounts for fruit softening. The aim of this work was to correlate molecular weight changes of WSF and OSF with the gene expression of pectin-solubilizing enzymes (pectinases) during papaya ripening. Papaya fruits obtained from a producer were harvest and storage under specific conditions. The fruits were divided in five groups according to days after harvesting. Cell walls from all groups of papaya pulp were isolated and fractionated (WSF and OSF). Expression profiles of pectinase genes were achieved according to the MIQE guidelines (Minimum Information for publication of Quantitative real-time PCR Experiments). The results showed an increased yield and a decreased molecular weight throughout ripening for WSF and OSF. Gene expression data support that papaya softening is achieved by polygalacturonases (PGs) up-regulation, in which their actions might have been facilitated by the constant action of pectinesterases (PMEs). Moreover, BGAL1 gene was up-regulated during ripening with a simultaneous galactose release, suggesting that galactosidases (GALs) could also account for pulp softening. The data suggest that a solubilization of galacturonans and a depolymerization of cell wall components were caused mainly by the action of PGs and GALs.

Keywords: carica papaya, fruit ripening, galactosidases, plant cell wall, polygalacturonases

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Authors: Jafar Ahmadi, Zhohreh Asiaban, Sedigheh Fabriki Ourang

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Brassinosteroids (BRs) regulate cell elongation, vascular differentiation, senescence and stress responses. BRs signal through the BES1/BZR1 family of transcription factors, which regulate hundreds of target genes involved in this pathway. In this research a comprehensive genome-wide analysis was carried out in BES1/BZR1 gene family in Arabidopsis thaliana, Cucumis sativus, Vitis vinifera, Glycin max, and Brachypodium distachyon. Specifications of the desired sequences, dot plot and hydropathy plot were analyzed in the protein and genome sequences of five plant species. The maximum amino acid length was attributed to protein sequence Brdic3g with 374aa and the minimum amino acid length was attributed to protein sequence Gm7g with 163aa. The maximum Instability index was attributed to protein sequence AT1G19350 equal with 79.99 and the minimum Instability index was attributed to protein sequence Gm5g equal with 33.22. Aliphatic index of these protein sequences ranged from 47.82 to 78.79 in Arabidopsis thaliana, 49.91 to 57.50 in Vitis vinifera, 55.09 to 82.43 in Glycin max, 54.09 to 54.28 in Brachypodium distachyon 55.36 to 56.83 in Cucumis sativus. Overall, data obtained from our investigation contributes a better understanding of the complexity of the BES1/BZR1 gene family and provides the first step towards directing future experimental designs to perform systematic analysis of the functions of the BES1/BZR1 gene family.

Keywords: BES1/BZR1, brassinosteroids, phylogenetic analysis, transcription factor

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Authors: Ceyda Okudu, Secil Eroglu, Khandakar A. S. M. Saadat, Sibel O. Balci

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Ring Finger Protein 2 (RNF2) is a member of polycomb repressive complex 1 (PRC1), which is one of the epigenetic regulators in the genome. When RNF2 combines with other PRC1 members, it mediates the mono-ubiquitination of Histon2A (H2A). In breast cancer, RNF2 is commonly overexpressed, and also it promotes metastasis and invasion in other aggressive tumors like melanoma, prostate, and hepatocarcinoma. The role of RNF2 in the metastasis and invasion of breast cancer has not yet been elucidated. Our aim is to observe the role of RNF2 in metastasis and invasion in this study by miRNA mediated RNF2 gene silencing in breast cancer cell lines. We selected miRNAs, targeting to RNF2 by searching online databases. miR-17-5p, miR20a-5p, and miR-106b-5p were transfected to breast cancer cell lines (MCF-7, MDA-MB-231, SK-BR-3, and ZR-75-1), and also we used normal breast epithelial cell line (hTERT-HME1) to compare RNF2 gene expression level. After 48-72 hours post-transfection, mRNAs were isolated from the cells, and gene expressions were measured by RT-qPCR after from cDNA syntheses. We observed that RNF2 was highly expressed in SK-BR-3 and MDA-MB-231 cell lines opposite to MCF-7 and ZR-75-1 cell lines. RNF2 was downregulated 5, 5 and 7 fold by miR17-5p, miR20a-5p and miR106b-5p respectively in MCF-7. However, in SK-BR-3 and ZR-75-1 cell lines, miRNAs did not affect significantly RNF2 gene expression level. miR20a-5p decreased RNF2 3 fold and miR17-5p and miR106b-5p did not affect MDA-MB-231. After gene expression analysis, we performed metastasis and invasion assay in MCF-7 cells. For metastasis, we used both wound healing assay and Transwell Cell Migration Assay, and we used Transwell Cell Invasion Assay for invasion. The data of this assay showed that miR17-5p and miR20a-5p decreased both invasion and metastasis level, but miR106b-5p has no effect. We would like to conclude that RNF2 can be targeted by miR17-5p, miR20a-5p and miR106b-5p in MCF-7 cells and also RNF2, which is one of the upregulated genes in aggressive tumor, can be decreased by using these miRNAs. In future, we would like to confirm these results at the protein level and also whether these miRNAs are direct target of RNF2 or not.

Keywords: breast cancer, epigenetic, microRNAs, RNF2

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Authors: Ebrahim Akbarzadeh, Josep A. Picas Barrachina, Maite Baile Puig

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This study focuses on a novel method for dispersion and distribution of reinforcement under high intensive shear stress to produce metal composites. The polyacrylonitrile (PAN)-based short carbon fiber (Csf) and Nextel 610 alumina fiber were dispersed under high intensive shearing at mushy zone in semi-solid of A356 by a novel method. The bundles and clusters were embedded by infiltration of slurry into the clusters, thus leading to a uniform microstructure. The fibers were embedded homogenously into the aluminum around 576-580°C with around 46% of solid fraction. Other experiments at 615°C and 568°C which are contained 0% and 90% solid respectively were not successful for dispersion and infiltration of aluminum into bundles of Csf. The alumina fiber has been cracked by high shearing load. The morphologies and crystalline phase were evaluated by SEM and XRD. The adopted thixo-process effectively improved the adherence and distribution of Csf into Al that can be developed to produce various composites by thixomixing.

Keywords: aluminum, carbon fiber, alumina fiber, thixomixing, adhesion

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Authors: Dana Gabriková, Martin Mistrík, Jarmila Bernasovská, Iveta Tóthová, Jana Kisková

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The Roma (Gypsies) is a transnational minority with a high degree of consanguineous marriages. Similar to other genetically isolated founder populations, the Roma harbor a number of unique or rare genetic disorders. This paper discusses about a rare form of Charcot-Marie-Tooth disease – type 4G (CMT4G), also called Hereditary Motor and Sensory Neuropathy type Russe, an autosomal recessive disease caused by mutation private to Roma characterized by abnormally increased density of non-myelinated axons. CMT4G was originally found in Bulgarian Roma and in 2009 two putative causative mutations in the HK1 gene were identified. Since then, several cases were reported in Roma families mainly from Bulgaria and Spain. Here we present a Slovak Roma family in which CMT4G was diagnosed on the basis of clinical examination and genetic testing. This case is a further proof of the role of the HK1 gene in pathogenesis of the disease. It confirms that mutation in the HK1 gene is a common cause of autosomal recessive CMT disease in Roma and should be considered as a common part of a diagnostic procedure.

Keywords: gypsies, HK1, HSMN-Russe, rare disease

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Authors: Ankan Roy, Niharika, Samir Kumar Patra

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Anomalous nexus of complex topological assemblies and spatiotemporal epigenetic choreography at chromosomal territory may forms the most sophisticated regulatory layer of gene expression in cancer. Colon cancer is one of the leading malignant neoplasms of the lower gastrointestinal tract worldwide. There is still a paucity of information about the complex molecular mechanisms of colonic cancerogenesis. Bioinformatics prediction and analysis helps to identify essential genes and significant pathways for monitoring and conquering this deadly disease. The present study investigates and explores potential hub genes as biomarkers and effective therapeutic targets for colon cancer treatment. Colon cancer patient sample containing gene expression profile datasets, such as GSE44076, GSE20916, and GSE37364 were downloaded from Gene Expression Omnibus (GEO) database and thoroughly screened using the GEO2R tool and Funrich software to find out common 2 differentially expressed genes (DEGs). Other approaches, including Gene Ontology (GO) and KEGG pathway analysis, Protein-Protein Interaction (PPI) network construction and hub gene investigation, Overall Survival (OS) analysis, gene correlation analysis, methylation pattern analysis, and hub gene-Transcription factors regulatory network construction, were performed and validated using various bioinformatics tool. Initially, we identified 166 DEGs, including 68 up-regulated and 98 down-regulated genes. Up-regulated genes are mainly associated with the Cytokine-cytokine receptor interaction, IL17 signaling pathway, ECM-receptor interaction, Focal adhesion and PI3K-Akt pathway. Downregulated genes are enriched in metabolic pathways, retinol metabolism, Steroid hormone biosynthesis, and bile secretion. From the protein-protein interaction network, thirty hub genes with high connectivity are selected using the MCODE and cytoHubba plugin. Survival analysis, expression validation, correlation analysis, and methylation pattern analysis were further verified using TCGA data. Finally, we predicted COL1A1, COL1A2, COL4A1, SPP1, SPARC, and THBS2 as potential master regulators in colonic cancerogenesis. Moreover, our experimental data highlights that disruption of lipid raft and RAS/MAPK signaling cascade affects this gene hub at mRNA level. We identified COL1A1, COL1A2, COL4A1, SPP1, SPARC, and THBS2 as determinant hub genes in colon cancer progression. They can be considered as biomarkers for diagnosis and promising therapeutic targets in colon cancer treatment. Additionally, our experimental data advertise that signaling pathway act as connecting link between membrane hub and gene hub.

Keywords: hub genes, colon cancer, DNA methylation, epigenetic engineering, bioinformatic predictions

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Authors: Monika Chuchro

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This paper presents a preliminary attempt to apply classification of time series using meta-clusters in order to improve the quality of regression models. In this case, clustering was performed as a method to obtain a subgroups of time series data with normal distribution from inflow into waste water treatment plant data which Composed of several groups differing by mean value. Two simple algorithms: K-mean and EM were chosen as a clustering method. The rand index was used to measure the similarity. After simple meta-clustering, regression model was performed for each subgroups. The final model was a sum of subgroups models. The quality of obtained model was compared with the regression model made using the same explanatory variables but with no clustering of data. Results were compared by determination coefficient (R2), measure of prediction accuracy mean absolute percentage error (MAPE) and comparison on linear chart. Preliminary results allows to foresee the potential of the presented technique.

Keywords: clustering, data analysis, data mining, predictive models

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Authors: Yun-Chin Chung, Nileema Divate, Gen-Hung Chen, Pei-Ru Huang, Rupesh Divate

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Many environmental factors, such as glucose concentration, ethanol, temperature, osmotic pressure and pH, decrease the production rate of ethanol using yeast as a starter. Fermentation starters with high tolerance to various stresses are always demanded for brewing industry. Trehalose, a storage carbohydrate in cell wall of yeast, plays an important role in tolerance of environmental stress by preserving integrity of plasma membrane and stabilizing proteins. Furan aldehydes are toxic to yeast and the growth rate of yeast is significantly reduced if furan aldehydes were present in the fermentation medium. In yeast, aldehyde reductase is involved in the detoxification of reactive aldehydes and consequently the growth of yeast is improved. The aims of this study were to construct a genetic recombinant Saccharomyces cerevisiae or Pichia pastoris with furfural and HMF degrading and high ethanol tolerance capacities. Yeast strains were engineered by genetic recombination for overexpression of trehalose-6-phosphate synthase gene (tps1) and aldehyde reductase gene (ari1). TPS1 gene was cloned from S. cerevisiae by reverse transcription-polymerase chain reaction (RT-PCR) and then ligated with pGAPZαC vector. The constructed vector, pGAPZC-tps1, was transformed to recombinant yeasts strain with overexpression of ari1. The transformants with pGAPZC-tps1-ari1 were generated called STA (S. cerevisiae) and PTA (P. pastoris) with overexpression of tps1, ari1. PCR with tps1-specific primers and western blot with his-tag confirmed the gene insertion and protein expression of tps1 in the transformants, respectively. The neutral trehalase gene (nth1) of STA was successfully deleted and the novel strain STAΔN will be used for further study, including the measurement of trehalose concentration and ethanol, furfural tolerance assay.

Keywords: genetic recombinant, yeast, ethanol tolerance, trehalase, aldehyde reductase

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Authors: Ali Bayram, Remzi Yiğiter

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Objective: Alzheimer's disease (AD), the most common cause of dementia, is a progressive neurodegenerative disease. At present, diagnosis of AD is rather late in the disease. Therefore, we attempted to find peripheral biomarkers for the early diagnosis of AD. Herein, we conducted a study to investigate the unc-51 like autophagy activating kinase-1 (ULK1) mRNA expression levels in human peripheral blood mononuclear cells from patients with Alzheimer's disease. Method: To determine whether ULK1 gene expression are altered in AD patients, we measured their gene expression in human peripheral blood cell in 50 patients with AD and 50 age and gender matched healthy controls by quantitative real-time PCR technique. Results: We found that both ULK1 gene expression in peripheral blood cell were significantly decreased in patients with AD as compared with controls (p <0.05). Lower levels of ULK1 gene expression were significantly associated with the increased risk for AD. Conclusions: Serine/threonine-protein kinase involved in autophagy in response to starvation. Acts upstream of phosphatidylinositol 3-kinase PIK3C3 to regulate the formation of autophagophores, the precursors of autophagosomes. Part of regulatory feedback loops in autophagy: acts both as a downstream effector and negative regulator of mammalian target of rapamycin complex 1 (mTORC1) via interaction with RPTOR. Activated via phosphorylation by AMPK and also acts as a regulator of AMPK by mediating phosphorylation of AMPK subunits PRKAA1, PRKAB2, and PRKAG1, leading to negatively regulate AMPK activity. May phosphorylate ATG13/KIAA0652 and RPTOR; however such data need additional evidences. Plays a role early in neuronal differentiation and is required for granule cell axon formation. Alzheimer is the most common neurodegenerative disease. Our results provide useful information that the ULK1 gene expression is decreased in the neurodegeneration and AD patients with, indicating their possible systemic involvement in AD.

Keywords: Alzheimer’s sisease, ULK1, mRNA expression, RT-PCR

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Authors: Jae Woong Sull, Sun Ha Jee

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High-density lipoprotein (HDL) cholesterol levels are associated with decreased risk of coronary artery disease. Several genome-wide association studies (GWAS) for HDL cholesterol levels have implicated cholesterol ester transfer protein (CETP) as possibly causal. We tested for the association between single nucleotide polymorphisms (SNPs) in CETP gene and HDL cholesterol levels in Korean population. Subjects were selected from the Korean Metabolic Syndrome Research Initiative study in the Bundang-Gu area. A total of 2,304 individuals from Bundang-Gu were recruited in 2008. Other subjects were selected from the Severance Hospital (N=4,294). SNP rs6499861 in the CETP gene was associated with mean HDL cholesterol levels (effect per allele -2.044 mg/dL, p=7.23×10-7). Subjects with the CG/GG genotype had a 1.46 -fold (range 1.24–1.72-fold) higher risk of having abnormal HDL cholesterol levels (<40 mg/dL) than subjects with the CC genotype. When analyzed by gender, the association of CETP was stronger in women than in men. When analyzed by physical activity behavior, the association with CETP was much stronger in male subjects with low physical activity (OR=1.54, 95% CI: 1.23-1.92, P=0.0001) than in male subjects with high physical activity. This study clearly demonstrates that genetic variants in CETP influence HDL cholesterol levels in Korean adults.

Keywords: CETP, HDL cholesterol, physical activity, polymorphisms

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Authors: Dipanshu Sur, Ratnabali Chakravorty

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Background: Polycystic ovary syndrome (PCOS) is the most common metabolic abnormality such as changes in lipid profile, diabetes, hypertension and metabolic syndrome occurring in young women of reproductive age. Low vitamin D levels were found to be associated with the development of obesity and insulin resistance in women with PCOS. Variants on vitamin D receptor (VDR) gene have also been related to metabolic comorbidities in general population. Aim: The aim of this case-control study was to investigate whether the VDR gene polymorphisms are associated with susceptibility to PCOS. Methods: Women with PCOS and a control group, all aged 16-40 years, were enrolled. Genotyping of VDR Fok-I (rs2228570), VDR Apa-I (rs7975232) as well as GC (rs2282679), DHCR7 (rs12785878) SNPs between groups were determined by using direct sequencing. Serum 25-hydroxyvitamin D [25(OH)] levels were measured by ELISA. Results: Mean serum 25(OH)D in the PCOS and control samples were 19.08±7 and 23.27±6.03 (p=0.048) which were significantly lower in PCOS patients compared with controls. CC genotype of the VDR Apa-I SNP was same frequent in PCOS (25.6%) and controls (25.6%) (OR: 0.9995; 95%CI: 0.528 to 1.8921; p= 0.9987). The CC genotype was also significantly associated with both lower E2 (p=0.031) and Androstenedione levels (p=0.062). We observed a significant association of GC polymorphism with 25(OH)D levels. PCOS women carrying the GG genotype (in GC genes) had significantly higher risk for vitamin D deficiency than women carrying the TT genotype. Conclusions: In conclusion, data from this study indicate that vitamin D levels are lower, and vitamin D deficiency more frequent, in PCOS than in controls. The present findings suggest that the Apa-I, Fok-I polymorphism of the VDR gene is associated with PCOS and seems to modulate ovarian steroid secretion. Further studies are needed to better clarify the biological mechanisms by which the polymorphism influences PCOS risk.

Keywords: vitamin D receptor, polymorphism, vitamin D, polycystic ovary syndrome

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Authors: Micheale Yifter Weldemichael, Hailay Mehari Gebremedhn, Teklehaimanot Hailesslasie

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The advancement of target-specific genome editing tools, including clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein9 (Cas9), mega-nucleases, base editing (BE), prime editing (PE), transcription activator-like endonucleases (TALENs), and zinc-finger nucleases (ZFNs), have paved the way for a modern era of gene editing. CRISPR/Cas9, as a versatile, simple, cost-effective and robust system for genome editing, has dominated the genome manipulation field over the last few years. The application of CRISPR/Cas9 in sorghum improvement is particularly vital in the context of ecological, environmental and agricultural challenges, as well as global climate change. In this context, gene editing using CRISPR/Cas9 can improve nutritional value, yield, resistance to pests and disease and tolerance to different abiotic stress. Moreover, CRISPR/Cas9 can potentially perform complex editing to reshape already available elite varieties and new genetic variations. However, existing research is targeted at improving even further the effectiveness of the CRISPR/Cas9 genome editing techniques to fruitfully edit endogenous sorghum genes. These findings suggest that genome editing is a feasible and successful venture in sorghum. Newer improvements and developments of CRISPR/Cas9 techniques have further qualified researchers to modify extra genes in sorghum with improved efficiency. The fruitful application and development of CRISPR techniques for genome editing in sorghum will not only help in gene discovery, creating new, improved traits in sorghum regulating gene expression sorghum functional genomics, but also in making site-specific integration events.

Keywords: CRISPR/Cas9, genome editing, quality, sorghum, stress, yield

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Authors: Philna Coetzee, Clara Msiza

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In modern society, audit committees are seen as the custodians of accountability and the conscience of management and the board. But who holds the audit committee accountable for their actions or non-actions and how do we know what they are supposed to be doing and what they are doing? The purpose of this article is to provide greater insight into the latter part of this problem, namely, determine what best practises for audit committees and the disclosure of what is the realities are. In countries where governance is well established, the roles and responsibilities of the audit committee are mostly clearly guided by legislation and/or guidance documents, with countries increasingly providing guidance on this topic. With high cost involved to adhere to governance guidelines, the public (for public organisations) and shareholders (for private organisations) expect to see the value of their ‘investment’. For audit committees, the dividends on the investment should reflect in less fraudulent activities, less corruption, higher efficiency and effectiveness, improved social and environmental impact, and increased profits, to name a few. If this is not the case (which is reflected in the number of fraudulent activities in both the private and the public sector), stakeholders have the right to ask: where was the audit committee? Therefore, the objective of this article is to contribute to the body of knowledge by comparing the adherence of audit committee to best practices guidelines as stipulated in the King Report across public listed companies, national and provincial government departments, state-owned enterprises and local municipalities. After constructs were formed, based on the literature, factor analyses were conducted to reduce the number of variables in each construct. Thereafter, cluster analyses, which is an explorative analysis technique that classifies a set of objects in such a way that objects that are more similar are grouped into the same group, were conducted. The SPSS TwoStep Clustering Component was used, being capable of handling both continuous and categorical variables. In the first step, a pre-clustering procedure clusters the objects into small sub-clusters, after which it clusters these sub-clusters into the desired number of clusters. The cluster analyses were conducted for each construct and the measure, namely the audit opinion as listed in the external audit report, were included. Analysing 228 organisations' information, the results indicate that there is a clear distinction between the four spheres of business that has been included in the analyses, indicating certain strengths and certain weaknesses within each sphere. The results may provide the overseers of audit committees’ insight into where a specific sector’s strengths and weaknesses lie. Audit committee chairs will be able to improve the areas where their audit committee is lacking behind. The strengthening of audit committees should result in an improvement of the accountability of boards, leading to less fraud and corruption.

Keywords: audit committee disclosure, cluster analyses, governance best practices, strengths and weaknesses

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Authors: Abdullah M. Alzahrani

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Our physiopathological assumption is that u-PA, t-PA, and PAI-1 are released by calcified aortic valves and play a role in the calcification of these valves. Sixty-five calcified aortic valves were collected from patients suffering from aortic stenosis. Each valve was incubated for 24 hours in culture medium. The supernatants were used to measure u-PA, t-PA, and PAI-1 concentrations; the valve calcification was evaluated using biphotonic absorptiometry. Aortic stenosis valves expressed normal plasminogen activators concentrations and overexpressed PAI-1 (u-PA, t-PA, and PAI-1 mean concentrations were, resp., 1.69 ng/mL ± 0.80, 2.76 ng/mL ± 1.33, and 53.27 ng/mL ± 36.39). There was no correlation between u-PA and PAI-1 (r = 0.3) but t-PA and PAI-1 were strongly correlated with each other (r = 0.6). Over expression of PAI-1 was proportional to the calcium content of theAS valves. Our results demonstrate a consistent increase of PAI-1 proportional to the calcification. The over expression of PAI-1 may be useful as a predictive indicator in patients with aortic stenosis.

Keywords: aortic valve, PAI-1, tPA gene, uPA gene

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Authors: Mustahsan Mir, Ahmed Hassanin, Mohammed A. Al-Saleh

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The layout optimization of building blocks of unequal areas has applications in many disciplines including VLSI floorplanning, macrocell placement, unequal-area facilities layout optimization, and plant or machine layout design. A number of heuristics and some analytical and hybrid techniques have been published to solve this problem. This paper presents an efficient high-quality building-block layout design technique especially suited for solving large-size problems. The higher efficiency and improved quality of optimized solutions are made possible by introducing the concept of Promising Infant Clusters in a constructive placement procedure. The results presented in the paper demonstrate the improved performance of the presented technique for benchmark problems in comparison with published heuristic, analytic, and hybrid techniques.

Keywords: block layout problem, building-block layout design, CAD, optimization, search techniques

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Authors: Omneya M. Helmy, Mona T. Kashef

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Aminoglycosides are used in treating a wide range of infections caused by both Gram-negative and Gram positive bacteria. The presence of 16S rRNA methyl transferases (16S-RMTase) is among the newly discovered resistance mechanisms that confer high resistance to clinically useful aminoglycosides. Cephalosporins are the most commonly used antimicrobials in Egypt; therefore, this study was conducted to determine the isolation frequency of 16S rRNA methyl transferases among third generation cephalosporin-resistant clinical isolates in Egypt. One hundred and twenty three cephalosporin resistant Gram-negative clinical isolates were screened for aminoglycoside resistance by the Kirby Bauer disk diffusion method and tested for possible production of 16S-RMTase. PCR testing and sequencing were used to confirm the presence of 16S-RMTase and the associated antimicrobial resistance determinants, as well as the genetic region surrounding the armA gene. Out of 123 isolates, 66 (53.66%) were resistant to at least one aminoglycoside antibiotic. Only one Escherichia coli isolate (E9ECMO) which was totally resistant to all tested aminoglycosides, was confirmed to have the armA gene in association with blaTEM-1, blaCTX-M-15, blaCTX-M-14 and aac(6)-Ib genes. The armA gene was found to be carried on a large A/C plasmid. Genetic mapping of the armA surrounding region revealed, for the first time, the association of armA with aac(6)-Ib on the same transposon. In Conclusion, the isolation frequency of 16S-RMTase was low among the tested cephalosporin-resistant clinical samples. However, a novel composite transposon has been detected conferring high-level aminoglycosides resistance.

Keywords: aminoglcosides, armA gene, β lactmases, 16S rRNA methyl transferases

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Authors: İsmail İnce, Mustafa Fener, Sair Kahraman

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The uniaxial compressive strength (UCS) of rocks is an important input parameter for the design of rock engineering project. Compressive strength can be determined in the laboratory using the uniaxial compressive strength (UCS) test. Although the test is relatively simple, the method is time consuming and expensive. Therefore many researchers have tried to assess the uniaxial compressive strength values of rocks via relatively simple and indirect tests (e.g. point load strength test, Schmidt Hammer hardness rebound test, P-wave velocity test, etc.). Pyroclastic rocks are widely exposed in the various regions of the world. Cappadocia region located in the Central Anatolia is one of the most spectacular cite of these regions. It is important to determine the mechanical behaviour of the pyroclastic rocks due to their ease of carving, heat insulation properties and building some civil engineering constructions in them. The purpose of this study is to estimate a widely varying uniaxial strength of pyroclastic rocks from Cappadocia region by means of point load strength, porosity, dry density and saturated density tests utilizing gene expression programming.

Keywords: pyroclastic rocks, uniaxial compressive strength, gene expression programming (GEP, Cappadocia region

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Authors: Carlo Stephen O. Moneva, Sharon Rose M. Tabugo

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The present study aims to assess polymorphism in the prolactin (C576A) gene and determine the influence of different prolactin (PRL) genotypes to milk yield performance in crossbred Anglo-Nubian dairy goats raised from Awang, Opol, Misamis Oriental and Talay, Dumaguete City, Negros Oriental. Genomic DNA was extracted from hair follicles and Polymerase Chain Reaction – Restriction Fragment Length Polymorphism (PCR-RFLP) was performed for the genotyping of the C576A polymorphism located in exon 5 of goats’ prolactin gene using Eco241 restriction enzyme. Genotypic and allelic frequencies of 0.56 for AA, 0.44 for AB, 0.78 for A, and 0.22 for B were recorded. Observed heterozygosity values were higher than the expected heterozygosity. All populations followed the Hardy–Weinberg principle at p>0.05, except for dairy goats from Farm A located in Opol, Misamis Oriental. A two-way factorial (2 x 4) in a Randomized Complete Block Design was used to be able to evaluate the relationship between genotypes and milk yield performance. PRL genotypes and parity were used as main factors and farm as the blocking factor. AB genotype goats produced significantly higher average daily milk yield and total milk production than AA genotype (p<0.05), an indication that the polymorphism in the caprine PRL (C576A) gene influenced milk yield performance in the population of crossbred Anglo-Nubian goats from Opol, Misamis Oriental and Dumaguete City, Negros Oriental. However, these results have to be validated in other dairy goat breeds.

Keywords: polymorphism, prolactin, milk yield, Anglo-Nubian, PCR-RFLP

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Authors: X. Cervantes-López, C. Arriaga-Canon, L. Contreras Espinosa

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The goal of this study is to validate the overexpression of the lncRNA GATA3-AS1 associated with resistance to neoadjuvant chemotherapy of female patients with locally advanced mammary adenocarcinoma of luminal B subtype This study involved a cohort of one hundred thirty-seven samples for which total RNA was isolated from formalin fixed paraffin embedded (FFPE) tissue. Samples were cut using a Microtome Hyrax M25 Zeiss and RNA was isolated using the RNeasy FFPE kit and a deparaffinization solution, the next step consisted in the analysis of RNA concentration and quality, then 18 µg of RNA was treated with DNase I, and cDNA was synthesized from 50 ng total RNA, finally real-time PCR was performed with SYBR Green/ROX qPCR Master Mix in order to determined relative gene expression using RPS28 as a housekeeping gene to normalize in a fold calculation ΔCt. As a result, we validated by real-time PCR that the overexpression of the lncRNA GATA3-AS1 is associated with resistance to neoadjuvant chemotherapy in locally advanced breast cancer patients of luminal B subtype.

Keywords: breast cancer, biomarkers, genomics, neoadjuvant chemotherapy, lncRNAS

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Authors: Mandeep Singh Azad.Dibyendu Chakraborty, Vikas Vohra

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Introduction: Poonch is one of the remotest districts of the Jammu and Kashmir (UT) and situated on international borders. This native poultry population in these areas is quite hardy and thrives well in adverse climatic conditions. Till date, no local breed from this area (Jammu Province) has been characterized thus present study was undertaken with the main objectives of molecular characterization of ChB6 gene in local native chicken of Poonch region located at international borders between India and Pakistan. The chicken B-cell marker (ChB6) gene has been proposed as a candidate gene in regulating B-cell development. Material and Method: RNA was isolated by Blood RNA Purification Kit (HiPura) and Trizol method from whole blood samples. Positive PCR products with size 1110 bp were selected for further purification, sequencing and analysis. The amplified PCR product was sequenced by Sangers dideoxy chain termination method. The obtained sequence of ChB6 gene of Poonchi chicken were compared by MEGAX software. BioEdit software was used to construct phylogenic tree, and Neighbor Joining method was used to infer evolutionary history. In order to compute evolutionary distance Maximum Composite Likelihood method was used. Results: The positively amplified samples of ChB6 genes were then subjected to Sanger sequencing with “Primer Walking. The sequences were then analyzed using MEGA X and BioEdit software. The sequence results were compared with other reported sequence from different breed of chicken and with other species obtained from the NCBI (National Center for Biotechnology Information). ClustalW method using MEGA X software was used for multiple sequence alignment. The sequence results of ChB6 gene of Poonchi chicken was compared with Centrocercus urophasianus, G. gallus mRNA for B6.1 protein, G. gallus mRNA for B6.2, G. gallus mRNA for B6.3, Gallus gallus B6.1, Halichoeres bivittatus, Miniopterus fuliginosus Ferringtonia patagonica, Tympanuchus phasianellus. The genetic distances were 0.2720, 0.0000, 0.0245, 0.0212, 0.0147, 1.6461, 2.2394, 2.0070 and 0.2363 for ChB6 gene of Poonchi chicken sequence with other sequences in the present study respectively. Sequencing results showed variations between different species. It was observed that AT content were higher then GC content for ChB6 gene. The lower AT content suggests less thermostable. It was observed that there was no sequence difference within the Poonchi population for ChB6 gene. The high homology within chicken population indicates the conservation of ChB6 gene. The maximum difference was observed with Miniopterus fuliginosus (Eastern bent-wing bat) followed by Ferringtonia patagonica and Halichoeres bivittatus. Conclusion: Genetic variation is the essential component for genetic improvement. The results of immune related gene Chb6 shows between population genetic variability. Therefore, further association studies of this gene with some prevalent diseases in large population would be helpful to identify disease resistant/ susceptible genotypes in the indigenous chicken population.

Keywords: ChB6, sequencing, ClustalW, genetic distance, poonchi chicken, SNP

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Authors: Ali Asheibi

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The rapid increase in the size of databases required to store power quality monitoring data has demanded new techniques for analysing and understanding the data. One suggested technique to assist in analysis is data mining. Preparing raw data to be ready for data mining exploration take up most of the effort and time spent in the whole data mining process. Clustering is an important technique in data mining and machine learning in which underlying and meaningful groups of data are discovered. Large amounts of harmonic data have been collected from an actual harmonic monitoring system in a distribution system in Australia for three years. This amount of acquired data makes it difficult to identify operational events that significantly impact the harmonics generated on the system. In this paper, harmonic data preparation processes to better understanding of the data have been presented. Underlying classes in this data has then been identified using clustering technique based on the Minimum Message Length (MML) method. The underlying operational information contained within the clusters can be rapidly visualised by the engineers. The C5.0 algorithm was used for classification and interpretation of the generated clusters.

Keywords: data mining, harmonic data, clustering, classification

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Authors: Mahmoud M. Zakaria, Omnia F. Elmoursi, Mahmoud M. Gabr, Camelia A. AbdelMalak, Mohamed A. Ghoneim

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Many protocols were publicized for differentiation of human mesenchymal stem cells (MSCS) into insulin-producing cells (IPCs) in order to excrete insulin hormone ingoing to treat diabetes disease. Our aim is to evaluate relative gene expression for each independent protocol. Human bone marrow cells were derived from three volunteers that suffer diabetes disease. After expansion of mesenchymal stem cells, differentiation of these cells was done by three different protocols (the one-step protocol was used conophylline protein, the two steps protocol was depending on trichostatin-A, and the three-step protocol was started by beta-mercaptoethanol). Evaluation of gene expression was carried out by real-time PCR: Pancreatic endocrine genes, transcription factors, glucose transporter, precursor markers, pancreatic enzymes, proteolytic cleavage, extracellular matrix and cell surface protein. Quantitation of insulin secretion was detected by immunofluorescence technique in 24-well plate. Most of the genes studied were up-regulated in the in vitro differentiated cells, and also insulin production was observed in the three independent protocols. There were some slight increases in expression of endocrine mRNA of two-step protocol and its insulin production. So, the two-step protocol was showed a more efficient in expressing of pancreatic endocrine genes and its insulin production than the other two protocols.

Keywords: mesenchymal stem cells, insulin producing cells, conophylline protein, trichostatin-A, beta-mercaptoethanol, gene expression, immunofluorescence technique

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Authors: A. Ferasat, S. Rostampour Yasouri

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Seasonal flu is a highly contagious infection caused by influenza viruses. These viruses undergo genetic changes that result in new epidemics across the globe. Medical attention is crucial in severe cases, particularly for the elderly, frail, and those with chronic illnesses, as their immune systems are often weaker. The purpose of this study was to detect new subtypes of the influenza A virus rapidly using a specific RT-PCR method based on the HA gene (hemagglutinin). In the winter and spring of 2022_2023, 120 embryonated egg samples were cultured, suspected of seasonal influenza. RNA synthesis, followed by cDNA synthesis, was performed. Finally, the PCR technique was applied using a pair of specific primers designed based on the HA gene. The PCR product was identified after purification, and the nucleotide sequence of purified PCR products was compared with the sequences in the gene bank. The results showed a high similarity between the sequence of the positive samples isolated from the patients and the sequence of the new strains isolated in recent years. This RT-PCR technique is entirely specific in this study, enabling the detection and multiplication of influenza and its subspecies from clinical samples. The RT-PCR technique based on the HA gene, along with sequencing, is a fast, specific, and sensitive diagnostic method for those infected with influenza viruses and its new subtypes. Rapid molecular diagnosis of influenza is essential for suspected people to control and prevent the spread of the disease to others. It also prevents the occurrence of secondary (sometimes fatal) pneumonia that results from influenza and pathogenic bacteria. The critical role of rapid diagnosis of new strains of influenza is to prepare a drug vaccine against the latest viruses that did not exist in the community last year and are entirely new viruses.

Keywords: influenza, molecular diagnosis, patients, RT-PCR technique

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Authors: R. Fardid, R. Coppes

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Introduction: In mediastinal radiotherapy and to a lesser extend also in total-body irradiation (TBI) radiation exposure may lead to development of cardiac diseases. Radiation-induced heart disease is dose-dependent and it is characterized by a loss of cardiac function, associated with progressive heart cells degeneration. We aimed to determine the in-vivo radiation effects on fibronectin, ColaA1, ColaA2, galectin and TGFb1 gene expression levels in left ventricle heart tissues of rats after irradiation. Material and method: Four non-treatment adult Wistar rats as control group (group A) were selected. In group B, 4 adult Wistar rats irradiated to 20 Gy single dose of 150 Mev proton beam locally in heart only. In heart plus lung irradiate group (group C) 4 adult rats was irradiated by 50% of lung laterally plus heart radiation that mentioned in before group. At 8 weeks after radiation animals sacrificed and left ventricle heart dropped in liquid nitrogen for RNA extraction by Absolutely RNA® Miniprep Kit (Stratagen, Cat no. 400800). cDNA was synthesized using M-MLV reverse transcriptase (Life Technologies, Cat no. 28025-013). We used Bio-Rad machine (Bio Rad iQ5 Real Time PCR) for QPCR testing by relative standard curve method. Results: We found that gene expression of fibronectin in group C significantly increased compared to control group, but it was not showed significant change in group B compared to group A. The levels of gene expressions of Cola1 and Cola2 in mRNA did not show any significant changes between normal and radiation groups. Changes of expression of galectin target significantly increased only in group C compared to group A. TGFb1 expressions in group C more than group B showed significant enhancement compared to group A. Conclusion: In summary we can say that 20 Gy of proton exposure of heart tissue may lead to detectable damages in heart cells and may distribute function of them as a component of heart tissue structure in molecular level.

Keywords: gene expression, heart damage, proton irradiation, radiotherapy

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Authors: Andi Aliah Hidayani, Asmi Citra Malina A. R. Tassakka, Andi Parenrengi

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Vanname Shrimp is one of the high yielding varieties that are more resistant to virus attacks. However, now this shrimp more death due to virus attack such as white spot disease caused by white spot syndrome virus (WSSV). Various efforts have done to prevent the disease, like immunostimulatory, probiotics, and vaccine. White spot syndrome virus (WSSV) envelope protein VP19 gene is important because of its involvement in the system infection of shrimp. This study aimed to isolate and characterize an envelope protein VP19 – encoding gene of WSSV using WSSV infected Vanname Shrimp sample from some areas in South Sulawesi (Pangkep, Barru and Pinrang). The genomic of DNA were isolated from shrimp muscle using DTAB-CTAB method. Isolation of gene encoding envelope protein VP19 WSSV ws successfully performed with the results of the length of DNA fragment was 387 bp. The results of homology analysis using BLASTn homology suggested that these isolates genes from Barru, Pangkep and Pinrang have closest relationship with isolates from Mexican.

Keywords: vanname, shrimp, WSSV, viral protein 19

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Authors: Michael P. Mannino, Gerald W. Hart

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The majority of glucose metabolism proceeds through glycolytic pathways such as glycolysis or pentose phosphate pathway, however, about 5% is shunted through the hexosamine biosynthetic pathway, producing uridine diphosphate N-acetyl glucosamine (UDP-GlcNAc). This precursor can then be incorporated into complex oligosaccharides decorating the cell surface or remain as an intracellular post-translational-modification (PTM) of serine/threonine residues (O-GlcNAcylation, OGN), which has been identified on over 4,000 cytosolic or nuclear proteins. Intracellular OGN has major implications on cellularprocesses, typically by modulating protein localization, protein-protein interactions, protein degradation, and gene expression. Additionally, OGN is known to have an extensive cross-talk with phosphorylation, be in a competitive or cooperative manner. Unlike other PTMs there are only two cycling enzymes that are capable of adding or removing the GlcNAc moiety, O-linked N-aceytl glucosamine Transferase (OGT) and O-linked N-acetyl glucoamidase (OGA), respectively. The activity of OGT has been shown to be sensitive to cellular UDP-GlcNAc levels, even changing substrate affinity. Owing to this and that the concentration of UDP-GlcNAc is related to the metabolisms of glucose, amino acid, fatty acid, and nucleotides, O-GlcNAc is often referred to as a nutrient sensing rheostat. Indeed OGN is known to regulate several signaling pathways as a result of nutrient levels, such as insulin signaling. Dysregulation of OGN is associated with several disease states such as cancer, diabetes, and neurodegeneration. Improvements in glycomics over the past 10-15 years has significantly increased the OGT substrate pool, suggesting O-GlcNAc’s involvement in a wide variety of signaling pathways. However, O-GlcNAc’s role at the receptor level has only been identified in a case-by-case basis of known pathways. Examining the OGN of the plasma membrane (PM) may better focus our understanding of O-GlcNAc-effected signaling pathways. In this current study, PM fractions were isolated from several cell types via ultracentrifugation, followed by purification and MS/MS analysis in several cell lines. This process was repeated with or without OGT/OGA inhibitors or with increased/decreased glucose levels in media to ascertain the importance of OGN. Various pathways are followed up on in more detailed studies employing methods to localize OGN at the PM specifically.

Keywords: GlcNAc, nutrient sensitive, post-translational-modification, receptor

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