Search results for: differentially expressed genes
Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 2003

Search results for: differentially expressed genes

1913 Gene Expression Analysis for Corals / Zooxanthellae under High Seawater Temperature Stress

Authors: Haruka Ito, Toru Maruyama, Michihiro Ito, Chuya Shinzato, Hiroyuki Fujimura, Yoshikatsu Nakano, Shoichiro Suda, Sachiyo Aburatani, Haruko Takeyama

Abstract:

Clarifying symbiotic relationships is one of the most important theme for understanding the marine eco-system. Coral reef has been regarded as an important environmental resource. Coral holobiont composed by coral, symbiotic microalgae zooxanthellae, and bacteria have complexed relationship. Zooxanthellae mainly supply organic matter to the host corals through their photosynthetic activity. The symbiotic relationship is indispensable for corals but may easily collapses due to the rise of seawater temperature. However, the molecular mechanism how seawater temperature influences their relationships still remain unclear. In this study, the transcriptomic analysis has applied to elucidate the coral-zooxanthellae relationships under high seawater temperature stress. To observe reactions of corals and zooxanthellae against the rise of seawater temperature, meta-gene expression in coral have been analyzed. The branches from six different colonies of a stony coral, Acropora tenuis, were sampled at nine times by 2016 at two locations, Ishikawabaru and South of Sesoko Island, Okinawa, Japan. The mRNAs extracted from the branches including zooxanthellae were sequenced by illumina HiSeq. Gene Set Enrichment Analysis (GSEA) based on hyper geometric distribution was performed. The seawater temperature at 2016 summer was unusually high, which was caused by El Niño event, and the number of zooxanthellae in coral was decreased in August. GSEA derived the several specific genes expressed in A. tenuis under heat stress conditions. The upregulated genes under heat stress highly related with infection immunity. The downregulated genes significantly contained cell cycle related genes. Thu, it is considered that heat stress cause disorder in cell metabolism of A. tenuis, resulting in serious influence to coral holobiont.

Keywords: coral, symbiosis, thermal stress response, transcriptome analysis

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1912 Association of Antibiotics Resistance with Efflux Pumps Genes among Multidrug-Resistant Klebsiella pneumonia Recovered from Hospital Waste Water Effluents in Eastern Cape, South Africa

Authors: Okafor Joan, Nwodo Uchechukwu

Abstract:

Klebsiella pneumoniae (K. pneumoniae) is a significant pathogen responsible for opportunistic and nosocomial infection. One of the most significant antibiotic resistance mechanisms in K. pneumoniae isolates is efflux pumps. Our current study identified efflux genes (AcrAB, OqxAB, MacAB, and TolC) and regulatory genes (RamR and RarA) in multidrug-resistant (MDR) K. pneumoniae isolated from hospital effluents and investigated their relationship with antibiotic resistance. The sum of 145 K. pneumoniae isolates was established by PCR and screened for antibiotic susceptibility. PCR detected efflux pump genes, and their link with antibiotic resistance was statistically examined. However, 120 (83%) of the confirmed isolated were multidrug-resistant, with the largest percentage of resistance to ampicillin (88.3%) and the weakest rate of resistance to imipenem (5.5%). Resistance to the other antibiotics ranged from 11% to 76.6%. Molecular distribution tests show that AcrA, AcrB, MacA, oqxB oqxA, TolC, MacB were detected in 96.7%, 85%, 76.7%, 70.8%, 55.8%, 39.1%, and 29.1% respectively. However, 14.3% of the isolates harboured all seven genes screened. Efflux pump system AcrAB (83.2%) was the most commonly detected in K. pneumonia isolated across all the antibiotics class-tested. In addition, the frequencies of RamR and RarA were 46.2% and 31.4%, respectively. AcrAB and OqxAB efflux pump genes were significantly associated with fluoroquinolone, beta-lactam, carbapenem, and tetracycline resistance (p<0.05). The high rate of efflux genes in this study demonstrated that this resistance mechanism is the dominant way in K. pneumoniae isolates. Appropriate treatment must be used to reduce and tackle the burden of resistant Klebsiella pneumonia in hospital wastewater effluents.

Keywords: Klebsiella pneumonia, efflux pumps, regulatory genes, multidrug-resistant, hospital, PCR

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1911 Altered Gene Expression: Induction/Suppression of some Pathogenesis Related Protein Genes in an Egyptian Isolate of Potato Leafroll Virus (PLRV)

Authors: Dalia G. Aseel

Abstract:

The potato (Solanum tubersum, L.) has become one of the major vegetable crops in Egypt and all over the world. Potato leafroll virus(PLRV) was observed on potato plants collected from different governorates in Egypt. Three cultivars, Spunta, Diamont, and Cara, infected with PLRV were collected; RNA was extracted and subjected to Real-Time PCR using the coat protein gene primers. The results showed that the expression of the coat protein was 39.6-fold, 12.45-fold, and 47.43-fold, respectively, for Spunta, Diamont, and Cara cultivars. Differential Display Polymerase Chain Reaction (DD-PCR) using pathogenesis-related protein 1 (PR-1), β-1,3-glucanases (PR-2), chitinase (PR-3), peroxidase (POD), and polyphenol oxidase (PPO) forward primers for pathogenesis-related proteins (PR). The obtained data revealed different banding patterns depending on the viral type and the region of infection. Regarding PLRV, 58 up-regulated and 19 down-regulated genes were detected. Sequence analysis of the up-and down-regulated genes revealed that infected plants were observed in comparison with the healthy control. Sequence analysis of the up-regulated gene was performed, and the encoding sequence analysis showed that the obtained genes include: induced stolen tip protein. On the other hand, two down-regulated genes were identified: disease resistance RPP-like protein and non-specific lipid-transfer protein. In this study, the expressions of PR-1, PR-2, PR-3, POD, and PPO genes in the infected leaves of three potato cultivars were estimated by quantitative real-time PCR. We can conclude that the PLRV-infection of potato plants inhibited the expression of the five PR genes. On the contrary, infected leaves by PLRV elevated the expression of some defense genes. This interaction may also induce and/or suppress the expression of some genes responsible for the plant's defense mechanisms.

Keywords: PLRV, pathogenesis-related proteins (PRs), DD-PCR, sequence, real-time PCR

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1910 Methylation Profiling and Validation of Candidate Tissue-Specific Differentially Methylated Regions for Identification of Human Blood, Saliva, Semen and Vaginal Fluid and Its Application in Forensics

Authors: Meenu Joshi, Natalie Naidoo, Farzeen Kader

Abstract:

Identification of body fluids is an essential step in forensic investigation to aid in crime reconstruction. Tissue-specific differentially methylated regions (tDMRs) of the human genome can be targeted to be used as biomarkers to differentiate between body fluids. The present study was undertaken to establish the methylation status of potential tDMRs in blood, semen, saliva, and vaginal fluid by using methylation-specific PCR (MSP) and bisulfite sequencing (BS). The methylation statuses of 3 potential tDMRS in genes ZNF282, PTPRS, and HPCAL1 were analysed in 10 samples of each body fluid. With MSP analysis, the ZNF282, and PTPRS1 tDMR displayed semen-specific hypomethylation while HPCAL1 tDMR showed saliva-specific hypomethylation. With quantitative analysis by BS, the ZNF282 tDMR showed statistically significant difference in overall methylation between semen and all other body fluids as well as at individual CpG sites (p < 0.05). To evaluate the effect of environmental conditions on the stability of methylation profiles of the ZNF282 tDMR, five samples of each body fluid were subjected to five different forensic simulated conditions (dry at room temperature, wet in an exsiccator, outside on the ground, sprayed with alcohol, and sprayed with bleach) for 50 days. Vaginal fluid showed highest DNA recovery under all conditions while semen had least DNA quantity. Under outside on the ground condition, all body fluids except semen showed a decrease in methylation level; however, a significant decrease in methylation level was observed for saliva. A statistical significant difference was observed for saliva and semen (p < 0.05) for outside on the ground condition. No differences in methylation level were observed for the ZNF282 tDMR under all conditions for vaginal fluid samples. Thus, in the present study ZNF282 tDMR has been identified as a novel and stable semen-specific hypomethylation marker.

Keywords: body fluids, bisulphite sequencing, forensics, tDMRs, MSP

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1909 Expression Profiling of Chlorophyll Biosynthesis Pathways in Chlorophyll B-Lacking Mutants of Rice (Oryza sativa L.)

Authors: Khiem M. Nguyen, Ming C. Yang

Abstract:

Chloroplast pigments are extremely important during photosynthesis since they play essential roles in light absorption and energy transfer. Therefore, understanding the efficiency of chlorophyll (Chl) biosynthesis could facilitate enhancement in photo-assimilates accumulation, and ultimately, in crop yield. The Chl-deficient mutants have been used extensively to study the Chl biosynthetic pathways and the biogenesis of the photosynthetic apparatus. Rice (Oryza sativa L.) is one of the most leading food crops, serving as staple food for many parts of the world. To author’s best knowledge, Chl b–lacking rice has been found; however the molecular mechanism of Chl biosynthesis still remains unclear compared to wild-type rice. In this study, the ultrastructure analysis, photosynthetic properties, and transcriptome profile of wild-type rice (Norin No.8, N8) and its Chl b-lacking mutant (Chlorina 1, C1) were examined. The finding concluded that total Chl content and Chl b content in the C1 leaves were strongly reduced compared to N8 leaves, suggesting that reduction in the total Chl content contributes to leaf color variation at the physiological level. Plastid ultrastructure of C1 possessed abnormal thylakoid membranes with loss of starch granule, large number of vesicles, and numerous plastoglobuli. The C1 rice also exhibited thinner stacked grana, which was caused by a reduction in the number of thylakoid membranes per granum. Thus, the different Chl a/b ratio of C1 may reflect the abnormal plastid development and function. Transcriptional analysis identified 23 differentially expressed genes (DEGs) and 671 transcription factors (TFs) that were involved in Chl metabolism, chloroplast development, cell division, and photosynthesis. The transcriptome profile and DEGs revealed that the gene encoding PsbR (PSII core protein) was down-regulated, therefore suggesting that the lower in light-harvesting complex proteins are responsible for the lower photosynthetic capacity in C1. In addition, expression level of cell division protein (FtsZ) genes were significantly reduced in C1, causing chloroplast division defect. A total of 19 DEGs were identified based on KEGG pathway assignment involving Chl biosynthesis pathway. Among these DEGs, the GluTR gene was down-regulated, whereas the UROD, CPOX, and MgCH genes were up-regulated. Observation through qPCR suggested that later stages of Chl biosynthesis were enhanced in C1, whereas the early stages were inhibited. Plastid structure analysis together with transcriptomic analysis suggested that the Chl a/b ratio was amplified both by the reduction in Chl contents accumulation, owning to abnormal chloroplast development, and by the enhanced conversion of Chl b to Chl a. Moreover, the results indicated the same Chl-cycle pattern in the wild-type and C1 rice, indicating another Chl b degradation pathway. Furthermore, the results demonstrated that normal grana stacking, along with the absence of Chl b and greatly reduced levels of Chl a in C1, provide evidence to support the conclusion that other factors along with LHCII proteins are involved in grana stacking. The findings of this study provide insight into the molecular mechanisms that underlie different Chl a/b ratios in rice.

Keywords: Chl-deficient mutant, grana stacked, photosynthesis, RNA-Seq, transcriptomic analysis

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1908 Exploring the Strategy to Identify Seed-Specific Acyl-Hydrolases from Arabidopsis thaliana by Activity-Based Protein Profiling

Authors: M. Latha, Achintya K. Dolui, P. Vijayaraj

Abstract:

Vegetable oils mainly triacylglycerol (TAG) are an essential nutrient in the human diet as well as one of the major global commodity. There is a pressing need to enhance the yield of oil production to meet the world’s growing demand. Oil content is controlled by the balance between synthesis and breakdown in the cells. Several studies have established to increase the oil content by the overexpression of oil biosynthetic enzymes. Interestingly the significant oil accumulation was observed with impaired TAG hydrolysis. Unfortunately, the structural, as well as the biochemical properties of the lipase enzymes, is widely unknown, and so far, no candidate gene was identified in seeds except sugar-dependent1 (SDP1). Evidence has shown that SDP1directly responsible for initiation of oil breakdown in the seeds during germination. The present study is the identification of seed-specific acyl-hydrolases by activity based proteome profiling (ABPP) using Arabidopsis thaliana as a model system. The ABPP reveals that around 8 to 10 proteins having the serine hydrolase domain and are expressed during germination of Arabidopsis seed. The N-term sequencing, as well as LC-MS/MS analysis, was performed for the differentially expressed protein during germination. The coding region of the identified proteins was cloned, and lipases activity was assessed with purified recombinant protein. The enzyme assay was performed against various lipid substrates, and we have observed the acylhydrolase activity towards lysophosphatidylcholine and monoacylglycerol. Further, the functional characteristic of the identified protein will reveal the physiological significance the enzyme in oil accumulation.

Keywords: lipase, lipids, vegetable oil, triacylglycerol

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1907 Detection of PCD-Related Transcription Factors for Improving Salt Tolerance in Plant

Authors: A. Bahieldin, A. Atef, S. Edris, N. O. Gadalla, S. M. Hassan, M. A. Al-Kordy, A. M. Ramadan, A. S. M. Al- Hajar, F. M. El-Domyati

Abstract:

The idea of this work is based on a natural exciting phenomenon suggesting that suppression of genes related to the program cell death (or PCD) mechanism might help the plant cells to efficiently tolerate abiotic stresses. The scope of this work was the detection of PCD-related transcription factors (TFs) that might also be related to salt stress tolerance in plant. Two model plants, e.g., tobacco and Arabidopsis, were utilized in order to investigate this phenomenon. Occurrence of PCD was first proven by Evans blue staining and DNA laddering after tobacco leaf discs were treated with oxalic acid (OA) treatment (20 mM) for 24 h. A number of 31 TFs up regulated after 2 h and co-expressed with genes harboring PCD-related domains were detected via RNA-Seq analysis and annotation. These TFs were knocked down via virus induced gene silencing (VIGS), an RNA interference (RNAi) approach, and tested for their influence on triggering PCD machinery. Then, Arabidopsis SALK knocked out T-DNA insertion mutants in selected TFs analogs to those in tobacco were tested under salt stress (up to 250 mM NaCl) in order to detect the influence of different TFs on conferring salt tolerance in Arabidopsis. Involvement of a number of candidate abiotic-stress related TFs was investigated.

Keywords: VIGS, PCD, RNA-Seq, transcription factors

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1906 Pathway and Differential Gene Expression Studies for Colorectal Cancer

Authors: Ankita Shukla, Tiratha Raj Singh

Abstract:

Colorectal cancer (CRC) imposes serious mortality burden worldwide and it has been increasing for past consecutive years. Continuous efforts have been made so far to diagnose the disease condition and to identify the root cause for it. In this study, we performed the pathway level as well as the differential gene expression studies for CRC. We analyzed the gene expression profile GSE24514 from Gene Expression Omnibus (GEO) along with the gene pathways involved in the CRC. This analysis helps us to understand the behavior of the genes that have shown differential expression through their targeted pathways. Pathway analysis for the targeted genes covers the wider area which therefore decreases the possibility to miss the significant ones. This will prove to be beneficial to expose the ones that have not been given attention so far. Through this analysis, we attempt to understand the various neighboring genes that have close relationship to the targeted one and thus proved to be significantly controlling the CRC. It is anticipated that the identified hub and neighboring genes will provide new directions to look at the pathway level differently and will be crucial for the regulatory processes of the disease.

Keywords: mismatch repair, microsatellite instability, carcinogenesis, morbidity

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1905 The Phylogenetic Investigation of Candidate Genes Related to Type II Diabetes in Man and Other Species

Authors: Srijoni Banerjee

Abstract:

Sequences of some of the candidate genes (e.g., CPE, CDKAL1, GCKR, HSD11B1, IGF2BP2, IRS1, LPIN1, PKLR, TNF, PPARG) implicated in some of the complex disease, e.g. Type II diabetes in man has been compared with other species to investigate phylogenetic affinity. Based on mRNA sequence of these genes of 7 to 8 species, using bioinformatics tools Mega 5, Bioedit, Clustal W, distance matrix was obtained. Phylogenetic trees were obtained by NJ and UPGMA clustering methods. The results of the phylogenetic analyses show that of the species compared: Xenopus l., Danio r., Macaca m., Homo sapiens s., Rattus n., Mus m. and Gallus g., Bos taurus, both NJ and UPGMA clustering show close affinity between clustering of Homo sapiens s. (Man) with Rattus n. (Rat), Mus m. species for the candidate genes, except in case of Lipin1 gene. The results support the functional similarity of these genes in physiological and biochemical process involving man and mouse/rat. Therefore, in understanding the complex etiology and treatment of the complex disease mouse/rate model is the best laboratory choice for experimentation.

Keywords: phylogeny, candidate gene of type-2 diabetes, CPE, CDKAL1, GCKR, HSD11B1, IGF2BP2, IRS1, LPIN1, PKLR, TNF, PPARG

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1904 Genome-Wide Identification and Characterization of MLO Family Genes in Pumpkin (Cucurbita maxima Duch.)

Authors: Khin Thanda Win, Chunying Zhang, Sanghyeob Lee

Abstract:

Mildew resistance locus o (Mlo), a plant-specific gene family with seven-transmembrane (TM), plays an important role in plant resistance to powdery mildew (PM). PM caused by Podosphaera xanthii is a widespread plant disease and probably represents the major fungal threat for many Cucurbits. The recent Cucurbita maxima genome sequence data provides an opportunity to identify and characterize the MLO gene family in this species. Total twenty genes (designated CmaMLO1 through CmaMLO20) have been identified by using an in silico cloning method with the MLO gene sequences of Cucumis sativus, Cucumis melo, Citrullus lanatus and Cucurbita pepo as probes. These CmaMLOs were evenly distributed on 15 chromosomes of 20 C. maxima chromosomes without any obvious clustering. Multiple sequence alignment showed that the common structural features of MLO gene family, such as TM domains, a calmodulin-binding domain and 30 important amino acid residues for MLO function, were well conserved. Phylogenetic analysis of the CmaMLO genes and other plant species reveals seven different clades (I through VII) and only clade IV is specific to monocots (rice, barley, and wheat). Phylogenetic and structural analyses provided preliminary evidence that five genes belonged to clade V could be the susceptibility genes which may play the importance role in PM resistance. This study is the first comprehensive report on MLO genes in C. maxima to our knowledge. These findings will facilitate the functional analysis of the MLOs related to PM susceptibility and are valuable resources for the development of disease resistance in pumpkin.

Keywords: Mildew resistance locus o (Mlo), powdery mildew, phylogenetic relationship, susceptibility genes

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1903 Diversities, Antibiogram and Antibiotic Resistance Genes in Staphylococcus Species in Raw Meat from a Research Farm

Authors: Anthony Ayodeji Adegoke, Olayinka Ayobami Aiyegoro, Thor Axel Stenstrom

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A study to investigate the species diversities, antibiogram and antibiotic resistance genes in Staphylococcus species from raw meat and dairy products collected from an abattoir and a farm shop of a research institute in Irene, South Africa over a six-month period was conducted. Polymerase Chain Reaction was used to speciate the bacteria and to detect the presence and otherwise of resistance genes. Antibiotic susceptibility testing was performed by disk diffusion method on Mueller-Hinton agar according to the Clinical Laboratory Standards Institute standards. A total of twenty-six (26) antibiotics were used to determine the antibiotic susceptibility. S. xylosus was the predominant isolate with 30% total occurrence, followed by S. epidermis, S. aureus, S. saprophyticus and S. haemolyticus with 25%, 15%, 15%, and 10% abundance respectively. The isolates were resistant to ceftezidime, gentamycin, nalidixic acid, nortrafuration, ampicillin, penicillin, oxytetracycline, tetracycline, doxycycline, clindamycin and lincomycin. mecA genes was detected among the methicillin resistant Staphylococcus species (MRSS) but no vancomycin resistance genes (van A and van B) were detected in these isolates. The presence of MRSS and multidrug resistant Staphylococcus species in meat affirms the need to avoid consumption of partially cooked meat currently rampant in South Africa, to avoid the spread of difficult to control pathogens in epidemiological proportion.

Keywords: Staphylococcus species, antibiotics, antibiotic resistance genes, food products, methicillin resistance, mecA gene

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1902 Evaluating Gene-Gene Interaction among Nicotine Dependence Genes on the Risk of Oral Clefts

Authors: Mengying Wang, Dongjing Liu, Holger Schwender, Ping Wang, Hongping Zhu, Tao Wu, Terri H Beaty

Abstract:

Background: Maternal smoking is a recognized risk factor for nonsyndromic cleft lip with or without cleft palate (NSCL/P). It has been reported that the effect of maternal smoking on oral clefts is mediated through genes that influence nicotine dependence. The polymorphisms of cholinergic receptor nicotinic alpha (CHRNA) and beta (CHRNB) subunits genes have previously shown strong associations with nicotine dependence. Here, we attempted to investigate whether the above genes are associated with clefting risk through testing for potential gene-gene (G×G) and gene-environment (G×E) interaction. Methods: We selected 120 markers in 14 genes associated with nicotine dependence to conduct transmission disequilibrium tests among 806 Chinese NSCL/P case-parent trios ascertained in an international consortium which conducted a genome-wide association study (GWAS) of oral clefts. We applied Cordell’s method using “TRIO” package in R to explore G×G as well as G×E interaction involving environmental tobacco smoke (ETS) based on conditional logistic regression model. Results: while no SNP showed significant association with NSCL/P after Bonferroni correction, we found signals for G×G interaction between 10 pairs of SNPs in CHRNA3, CHRNA5, and CHRNB4 (p<10-8), among which the most significant interaction was found between RS3743077 (CHRNA3) and RS11636753 (CHRNB4, p<8.2×10-12). Linkage disequilibrium (LD) analysis revealed only low level of LD between these markers. However, there were no significant results for G×ETS interaction. Conclusion: This study fails to detect association between nicotine dependence genes and NSCL/P, but illustrates the importance of taking into account potential G×G interaction for genetic association analysis in NSCL/P. This study also suggests nicotine dependence genes should be considered as important candidate genes for NSCL/P in future studies.

Keywords: Gene-Gene Interaction, Maternal Smoking, Nicotine Dependence, Non-Syndromic Cleft Lip with or without Cleft Palate

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1901 Genomics Approach for Excavation of NAS Genes from Nutri Rich Minor Millet Crops: Transforming Perspective from Orphan Plants to Future Food Crops

Authors: Mahima Dubey, Girish Chandel

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Minor millets are highly nutritious and climate resilient cereal crops. These features make them ideal candidates to excavate the physiology of the underlying mechanism. In an attempt to understand the basis of mineral nutrition in minor millets, a set of five Barnyard millet genotypes were analyzed for grain Fe and Zn content under contrasting Fe-Zn supply to identify genotypes proficient in tolerating mineral deficiency. This resulted in the identification of Melghat-1 genotype to be nutritionally superior with better ability to withstand deficiency. Expression analysis of several Nicotianamine synthase (NAS) genes showed that HvNAS1 and OsNAS2 genes were prominent in positively mediating mineral deficiency response in Barnyard millet. Further, strategic efforts were employed for fast-track identification of more effective orthologous NAS genes from Barnyard millet. This resulted in the identification of two genes namely EfNAS1 (orthologous to HvNAS1 of barley) and EfNAS2 (orthologous to OsNAS2 gene of rice). Sequencing and thorough characterization of these sequences revealed the presence of intact NAS domain and signature tyrosine and di-leucine motifs in their predicted proteins and thus established their candidature as functional NAS genes in Barnyard millet. Moreover, EfNAS1 showed structural superiority over previously known NAS genes and is anticipated to have role in more efficient metal transport. Findings of the study provide insight into Fe-Zn deficiency response and mineral nutrition in millets. This provides millets with a physiological edge over micronutrient deficient staple cereals such as rice in withstanding Fe-Zn deficiency and subsequently accumulating higher levels of Fe and Zn in millet grains.

Keywords: gene expression, micronutrient, millet, ortholog

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1900 Investigation of Carbapenem-Resistant Genes in Acinetobacter spp. Isolated from Patients at Tertiary Health Care Center, Northeastern Thailand

Authors: S. J. Sirima, C. Thirawan, R.Puntharikorn, K. Ungsumalin, J. Kaemwich

Abstract:

Acinetobacter spp. is a gram negative bacterium causing the high incidence of multi-drug resistance in patients admitted to an intensive care unit. A hundred isolates of Imipenem-resistant Acinetobacter spp. isolated from patients admitted at tertiary health care center, Northeastern region, Ubon Ratchathani, Thailand, were subjected to modified Hodge test and combined disc test in order to evaluate the production of carbapenemases. The results revealed that about 35% of isolates were found to be carbapenemases producers. In addition, multiplex polymerase chain reactions were performed to detect blaOXA-like genes. It showed that 92% of isolates possess blaOXA-51-like and blaOXA-23-like genes. However, blaOXA-58-like gene was detected in only 8 isolates. No detection of blaOXA-24-like gene was observed in all isolates. In conclusion, an ability to produce carbepenemases would be an important mechanism of multi-drug resistance among clinical isolates of Acinetobacter spp. at tertiary health care center, Northeastern region, Ubon Ratchathani, Thailand. Furthermore, it was likely that the class D carbapenemases genes, blaOXA-51-like and blaOXA-23-like, might contribute to imipenem-resistance exhibiting among isolates.

Keywords: Acinetobacter spp., blaOXA-like genes, carbapenemases, tertiary health care center

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1899 Parkinson's Disease Gene Identification Using Physicochemical Properties of Amino Acids

Authors: Priya Arora, Ashutosh Mishra

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Gene identification, towards the pursuit of mutated genes, leading to Parkinson’s disease, puts forward a challenge towards proactive cure of the disorder itself. Computational analysis is an effective technique for exploring genes in the form of protein sequences, as the theoretical and manual analysis is infeasible. The limitations and effectiveness of a particular computational method are entirely dependent on the previous data that is available for disease identification. The article presents a sequence-based classification method for the identification of genes responsible for Parkinson’s disease. During the initiation phase, the physicochemical properties of amino acids transform protein sequences into a feature vector. The second phase of the method employs Jaccard distances to select negative genes from the candidate population. The third phase involves artificial neural networks for making final predictions. The proposed approach is compared with the state of art methods on the basis of F-measure. The results confirm and estimate the efficiency of the method.

Keywords: disease gene identification, Parkinson’s disease, physicochemical properties of amino acid, protein sequences

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1898 High-Risk Gene Variant Profiling Models Ethnic Disparities in Diabetes Vulnerability

Authors: Jianhua Zhang, Weiping Chen, Guanjie Chen, Jason Flannick, Emma Fikse, Glenda Smerin, Yanqin Yang, Yulong Li, John A. Hanover, William F. Simonds

Abstract:

Ethnic disparities in many diseases are well recognized and reflect the consequences of genetic, behavior, and environmental factors. However, direct scientific evidence connecting the ethnic genetic variations and the disease disparities has been elusive, which may have led to the ethnic inequalities in large scale genetic studies. Through the genome-wide analysis of data representing 185,934 subjects, including 14,955 from our own studies of the African America Diabetes Mellitus, we discovered sets of genetic variants either unique to or conserved in all ethnicities. We further developed a quantitative gene function-based high-risk variant index (hrVI) of 20,428 genes to establish profiles that strongly correlate with the subjects' self-identified ethnicities. With respect to the ability to detect human essential and pathogenic genes, the hrVI analysis method is both comparable with and complementary to the well-known genetic analysis methods, pLI and VIRlof. Application of the ethnicity-specific hrVI analysis to the type 2 diabetes mellitus (T2DM) national repository, containing 20,791 cases and 24,440 controls, identified 114 candidate T2DM-associated genes, 8.8-fold greater than that of ethnicity-blind analysis. All the genes identified are defined as either pathogenic or likely-pathogenic in ClinVar database, with 33.3% diabetes-associated and 54.4% obesity-associated genes. These results demonstrate the utility of hrVI analysis and provide the first genetic evidence by clustering patterns of how genetic variations among ethnicities may impede the discovery of diabetes and foreseeably other disease-associated genes.

Keywords: diabetes-associated genes, ethnic health disparities, high-risk variant index, hrVI, T2DM

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1897 Molecular Characterization of Ovine Herpesvirus 2 Strains Based on Selected Glycoprotein and Tegument Genes

Authors: Fulufhelo Amanda Doboro, Kgomotso Sebeko, Stephen Njiro, Moritz Van Vuuren

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Ovine herpesvirus 2 (OvHV-2) genome obtained from the lymphopblastoid cell line of a BJ1035 cow was recently sequenced in the United States of America (USA). Information on the sequences of OvHV-2 genes obtained from South African strains from bovine or other African countries and molecular characterization of OvHV-2 is not documented. Present investigation provides information on the nucleotide and derived amino acid sequences and genetic diversity of Ov 7, Ov 8 ex2, ORF 27 and ORF 73 genes, of these genes from OvHV-2 strains circulating in South Africa. Gene-specific primers were designed and used for PCR of DNA extracted from 42 bovine blood samples that previously tested positive for OvHV-2. The expected PCR products of 495 bp, 253 bp, 890 bp and 1632 bp respectively for Ov 7, Ov 8 ex2, ORF 27 and ORF 73 genes were sequenced and multiple sequence analysis done on the selected regions of the sequenced PCR products. Two genotypes for ORF 27 and ORF 73 gene sequences, and three genotypes for Ov 7 and Ov 8 ex2 gene sequences were identified, and similar groupings for the derived amino acid sequences were obtained for each gene. Nucleotide and amino acid sequence variations that led to the identification of the different genotypes included SNPs, deletions and insertions. Sequence analysis of Ov 7 and ORF 27 genes revealed variations that distinguished between sequences from SA and reference OvHV-2 strains. The implication of geographic origin among SA sequences was difficult to evaluate because of random distribution of genotypes in the different provinces, for each gene. However, socio-economic factors such as migration of people with animals, or transportation of animals for agricultural or business use from one province to another are most likely to be responsible for this observation. The sequence variations observed in this study have no impact on the antibody binding activities of glycoproteins encoded by Ov 7, Ov 8 ex2 and ORF 27 genes, as determined by prediction of the presence of B cell epitopes using BepiPred 1.0. The findings of this study will be used for selection of gene candidates for the development of diagnostic assays and vaccine development as well.

Keywords: amino acid, genetic diversity, genes, nucleotide

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1896 Genome-Wide Identification of Genes Resistance to Nitric Oxide in Vibrio parahaemolyticus

Authors: Yantao Li, Jun Zheng

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Food poison caused by consumption of contaminated food, especially seafood, is one of most serious public health threats worldwide. Vibrio parahaemolyticus is emerging bacterial pathogen and the leading cause of human gastroenteritis associated with food poison, especially in the southern coastal region of China. To successfully cause disease in host, bacterial pathogens need to overcome the host-derived stresses encountered during infection. One of the toxic chemical species elaborated by the host is nitric oxide (NO). NO is generated by acidified nitrite in the stomach and by enzymes of the inducible NO synthase (iNOS) in the host cell, and is toxic to bacteria. Bacterial pathogens have evolved some mechanisms to battle with this toxic stress. Such mechanisms include genes to sense NO produced from immune system and activate others to detoxify NO toxicity, and genes to repair the damage caused by toxic reactive nitrogen species (RNS) generated during NO toxic stress. However, little is known about the NO resistance in V. parahaemolyticus. In this study, a transposon coupled with next generation sequencing (Tn-seq) technology will be utilized to identify genes for NO resistance in V. parahaemolyticus. Our strategy will include construction the saturating transposon insertion library, transposon library challenging with NO, next generation sequencing (NGS), bioinformatics analysis and verification of the identified genes in vitro and in vivo.

Keywords: vibrio parahaemolyticus, nitric oxide, tn-seq, virulence

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1895 ScRNA-Seq RNA Sequencing-Based Program-Polygenic Risk Scores Associated with Pancreatic Cancer Risks in the UK Biobank Cohort

Authors: Yelin Zhao, Xinxiu Li, Martin Smelik, Oleg Sysoev, Firoj Mahmud, Dina Mansour Aly, Mikael Benson

Abstract:

Background: Early diagnosis of pancreatic cancer is clinically challenging due to vague, or no symptoms, and lack of biomarkers. Polygenic risk score (PRS) scores may provide a valuable tool to assess increased or decreased risk of PC. This study aimed to develop such PRS by filtering genetic variants identified by GWAS using transcriptional programs identified by single-cell RNA sequencing (scRNA-seq). Methods: ScRNA-seq data from 24 pancreatic ductal adenocarcinoma (PDAC) tumor samples and 11 normal pancreases were analyzed to identify differentially expressed genes (DEGs) in in tumor and microenvironment cell types compared to healthy tissues. Pathway analysis showed that the DEGs were enriched for hundreds of significant pathways. These were clustered into 40 “programs” based on gene similarity, using the Jaccard index. Published genetic variants associated with PDAC were mapped to each program to generate program PRSs (pPRSs). These pPRSs, along with five previously published PRSs (PGS000083, PGS000725, PGS000663, PGS000159, and PGS002264), were evaluated in a European-origin population from the UK Biobank, consisting of 1,310 PDAC participants and 407,473 non-pancreatic cancer participants. Stepwise Cox regression analysis was performed to determine associations between pPRSs with the development of PC, with adjustments of sex and principal components of genetic ancestry. Results: The PDAC genetic variants were mapped to 23 programs and were used to generate pPRSs for these programs. Four distinct pPRSs (P1, P6, P11, and P16) and two published PRSs (PGS000663 and PGS002264) were significantly associated with an increased risk of developing PC. Among these, P6 exhibited the greatest hazard ratio (adjusted HR[95% CI] = 1.67[1.14-2.45], p = 0.008). In contrast, P10 and P4 were associated with lower risk of developing PC (adjusted HR[95% CI] = 0.58[0.42-0.81], p = 0.001, and adjusted HR[95% CI] = 0.75[0.59-0.96], p = 0.019). By comparison, two of the five published PRS exhibited an association with PDAC onset with HR (PGS000663: adjusted HR[95% CI] = 1.24[1.14-1.35], p < 0.001 and PGS002264: adjusted HR[95% CI] = 1.14[1.07-1.22], p < 0.001). Conclusion: Compared to published PRSs, scRNA-seq-based pPRSs may be used not only to assess increased but also decreased risk of PDAC.

Keywords: cox regression, pancreatic cancer, polygenic risk score, scRNA-seq, UK biobank

Procedia PDF Downloads 55
1894 In Silico Analysis of Small Heat Shock Protein Gene Family by RNA-Seq during Tomato Fruit Ripening

Authors: Debora P. Arce, Flavia J. Krsticevic, Marco R. Bertolaccini, Joaquín Ezpeleta, Estela M. Valle, Sergio D. Ponce, Elizabeth Tapia

Abstract:

Small Heat Shock Proteins (sHSPs) are low molecular weight chaperones that play an important role during stress response and development in all living organisms. Fruit maturation and oxidative stress can induce sHSP synthesis both in Arabidopsis and tomato plants. RNA-Seq technology is becoming widely used in various transcriptomics studies; however, analyzing and interpreting the RNA-Seq data face serious challenges. In the present work, we de novo assembled the Solanum lycopersicum transcriptome for three different maturation stages (mature green, breaker and red ripe). Differential gene expression analysis was carried out during tomato fruit development. We identified 12 sHSPs differentially expressed that might be involved in breaker and red ripe fruit maturation. Interestingly, these sHSPs have different subcellular localization and suggest a complex regulation of the fruit maturation network process.

Keywords: sHSPs, maturation, tomato, RNA-Seq, assembly

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1893 Genomic Imprinting as a Possible Epigenetic Cause of Esophageal Atresia

Authors: M. Błoch, P. Karpiński, P. Gasperowicz, R. Płoski, A. Lebioda, P. Skiba, A. Rozensztrauch, D. Patkowski, R. Śmigiel

Abstract:

Introduction: The cause of the isolated form of esophageal atresia has been yet unknown. Objectives: The primary objective of this study was to indicate epigenetic factors which may play an important role in the etiopathogenesis of esophageal atresia. Methods: We recruited a group of 6 pairs of twins, among whom one of the twins developed EA. The selection of such a group for testing allows for excluding external factors (e.g., infections, drugs, toxins) as the cause of the birth defect. The analyzes were performed with the use of genetic material isolated from the whole blood and esophagus tissue of a patient with EA. The reduced representation bisulphite sequencing (RRBS) technique was used to study the change in the genomic imprinting -a change in the expression of genes, which may be the epigenetic cause of EA. Results: In the course of the analyzes, significant hypomethylation and hypermethylation regions were identified. 65 genes with probably increased expression and 65 with decreased expression were selected. These genes have not been marked in literature as possibly pathogenic in esophageal atresia. However, their participation in the pathogenesis of esophageal atresia cannot be clearly excluded. Conclusion: We suggest a role of hypomethylation or hypermethylation of selected genes as one of the possible epigenetic factors in EA pathogenesis. The use of the RRBS technique in the search for the cause of EA is pioneer research; therefore, it seems necessary to extend the research group to new patients with EA. Acknowledgment: The work was supported by the National Science Centre, Poland, under research project 2016/21/N/NZ5/01927.

Keywords: esophageal atresia, epigenetics, embryonic development, surgery, genes expression, twins

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1892 The Evaluation of Adjuvant Effects of CD154 in a Subunit Vaccine against Classical Swine Fever Virus

Authors: Yu-Chieh Chen, Li-Yun Wang, Chi-Chih Chen, Huy Hùng Đào, Ya-Mei Chen, Ming-Chu Cheng, Wen-Bin Chung, Hso-Chi Chaung, Guan-Ming Ke

Abstract:

Many recent researches have demonstrated that CD154, a protein primarily expressed on activated T cell molecules, has potentially acted as a molecular adjuvant to improve the immunogenicity of subunit vaccines against viral infections. Classical swine fever (CSF) affects the swine industry worldwide that is one of the most devastating and highly contagious pig diseases. It is listed by the World Organization for Animal Health (OIE) as an infectious animal disease that must be reported. Although pigs vaccinated with subunit vaccines can be differentially diagnosed from those infected animals, subunit vaccines usually need adjuvants to enhance and elicit immune responses. In this study, CD154 was linked with CSFV E2 sequences and then expressed in CHO cells to produce the fusion protein as E2-CD154. The porcine specific CpG adjuvant was also used in one of the formulations. The specific pathogen-free pigs (SPF) at the age of 4-week-old were randomly separated into four groups, vaccinated with E2-CpG, E2-CD154, E2-CD154-CpG or the commercial Bayovac® CSF-E2 vaccine and boosted two weeks after primary vaccination. The results showed that the percentages of CD4+ and CD4+IL2+ in peripheral blood mononuclear cells (PBMC) in E2-CD154 vaccinated piglets seven days after primary vaccination were gained by 1-5% relative to the control group. In addition, the percentages of CD4+IFNγ+ T cells had slightly edged up 0.1-0.3% compared with the control group. Also, increased E2-specific IFNγ levels had edged up CD4+CD8+ T cells found in E2-CD154 and E2-CD154-CpG groups, particularly in the E2-CD154-CpG group. These results implicate that CD154 may enhance cellular immunity and synergistically act with species-specific CpG adjuvant as a dual-phase adjuvant. Therefore, the CD154 may be beneficial as a promising adjuvant in subunit vaccines.

Keywords: CD154, CpG adjuvant, cellular immunity, subunit vaccine, pig

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1891 Application of KL Divergence for Estimation of Each Metabolic Pathway Genes

Authors: Shohei Maruyama, Yasuo Matsuyama, Sachiyo Aburatani

Abstract:

The development of the method to annotate unknown gene functions is an important task in bioinformatics. One of the approaches for the annotation is The identification of the metabolic pathway that genes are involved in. Gene expression data have been utilized for the identification, since gene expression data reflect various intracellular phenomena. However, it has been difficult to estimate the gene function with high accuracy. It is considered that the low accuracy of the estimation is caused by the difficulty of accurately measuring a gene expression. Even though they are measured under the same condition, the gene expressions will vary usually. In this study, we proposed a feature extraction method focusing on the variability of gene expressions to estimate the genes' metabolic pathway accurately. First, we estimated the distribution of each gene expression from replicate data. Next, we calculated the similarity between all gene pairs by KL divergence, which is a method for calculating the similarity between distributions. Finally, we utilized the similarity vectors as feature vectors and trained the multiclass SVM for identifying the genes' metabolic pathway. To evaluate our developed method, we applied the method to budding yeast and trained the multiclass SVM for identifying the seven metabolic pathways. As a result, the accuracy that calculated by our developed method was higher than the one that calculated from the raw gene expression data. Thus, our developed method combined with KL divergence is useful for identifying the genes' metabolic pathway.

Keywords: metabolic pathways, gene expression data, microarray, Kullback–Leibler divergence, KL divergence, support vector machines, SVM, machine learning

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1890 Melanoma Antigen Proteins Are Involved in DNA Damage Response

Authors: Olivier de Backer, Alexis Khelfi, Olivier Svensek, Axelle Nolmans, Dominique Desnoeck

Abstract:

The SMC5-SMC6 complex helps replication and repair of DNA double-strand breaks. Nse1, Nse3 and Nse4 are non-SMC components of the complex in which Nse3 stimulates the E3 ubiquitin ligase activity of Nse1 and is required for recruiting the complex on DNA. In most eukaryotes, Nse3 is a single protein, but in eutherians (placental mammals), it belongs to a large family of proteins called MAGE (Melanoma antigen) that share a conserved domain of about 200 aa known as MHD (Mage homology domain). MAGE assembles specific RING and HECT ubiquitin ligases and determines new substrates for ubiquitination. The MHD is required for the interaction with the cognate E3 ligase. Some MAGEs (referred to as Type I) are exclusively expressed in germ cells of the testis but are often expressed ectopically in cancer cells as the result of epigenetic modifications. The 12 MAGE-A genes belong to this category. Serval MAGE-A proteins could promote tumorigenesis by targeting tumor suppressor proteins (including p53) for ubiquitination and degradation. We showed that depletion of MAGE-A proteins in melanoma cells results in impaired DNA damage response and increased double-strand breaks after exposure to camptothecin. Moreover, it was shown that other actors of the DNA Damage Response were impacted when cells were depleted of MAGEA proteins.

Keywords: DNA damage response, melanoma, camptothecin, new role, MAGEA

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1889 Time-Course Lipid Accumulation and Transcript Analyses of Lipid Biosynthesis Gene of Chlorella sp.3 under Nitrogen Limited Condition

Authors: Jyoti Singh, Swati Dubey, Mukta Singh, R. P. Singh

Abstract:

The freshwater microalgae Chlorella sp. is alluring considerable interest as a source for biofuel production due to its fast growth rate and high lipid content. Under nitrogen limited conditions, they can accumulate significant amounts of lipids. Thus, it is important to gain insight into the molecular mechanism of their lipid metabolism. In this study under nitrogen limited conditions, regular pattern of growth characteristics lipid accumulation and gene expression analysis of key regulatory genes of lipid biosynthetic pathway were carried out in microalgae Chlorella sp 3. Our results indicated that under nitrogen limited conditions there is a significant increase in the lipid content and lipid productivity, achieving 44.21±2.64 % and 39.34±0.66 mg/l/d at the end of the cultivation, respectively. Time-course transcript patterns of lipid biosynthesis genes i.e. acetyl coA carboxylase (accD) and diacylglycerol acyltransferase (dgat) showed that during late log phase of microalgae Chlorella sp.3 both the genes were significantly up regulated as compared to early log phase. Moreover, the transcript level of the dgat gene is two-fold higher than the accD gene. The results suggested that both the genes responded sensitively to the nitrogen limited conditions during the late log stage, which proposed their close relevance to lipid biosynthesis. Further, this transcriptome data will be useful for engineering microalgae species by targeting these genes for genetic modification to improve microalgal biofuel quality and production.

Keywords: biofuel, gene, lipid, microalgae

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1888 Identification of New Familial Breast Cancer Susceptibility Genes: Are We There Yet?

Authors: Ian Campbell, Gillian Mitchell, Paul James, Na Li, Ella Thompson

Abstract:

The genetic cause of the majority of multiple-case breast cancer families remains unresolved. Next generation sequencing has emerged as an efficient strategy for identifying predisposing mutations in individuals with inherited cancer. We are conducting whole exome sequence analysis of germ line DNA from multiple affected relatives from breast cancer families, with the aim of identifying rare protein truncating and non-synonymous variants that are likely to include novel cancer predisposing mutations. Data from more than 200 exomes show that on average each individual carries 30-50 protein truncating mutations and 300-400 rare non-synonymous variants. Heterogeneity among our exome data strongly suggest that numerous moderate penetrance genes remain to be discovered, with each gene individually accounting for only a small fraction of families (~0.5%). This scenario marks validation of candidate breast cancer predisposing genes in large case-control studies as the rate-limiting step in resolving the missing heritability of breast cancer. The aim of this study is to screen genes that are recurrently mutated among our exome data in a larger cohort of cases and controls to assess the prevalence of inactivating mutations that may be associated with breast cancer risk. We are using the Agilent HaloPlex Target Enrichment System to screen the coding regions of 168 genes in 1,000 BRCA1/2 mutation-negative familial breast cancer cases and 1,000 cancer-naive controls. To date, our interim analysis has identified 21 genes which carry an excess of truncating mutations in multiple breast cancer families versus controls. Established breast cancer susceptibility gene PALB2 is the most frequently mutated gene (13/998 cases versus 0/1009 controls), but other interesting candidates include NPSR1, GSN, POLD2, and TOX3. These and other genes are being validated in a second cohort of 1,000 cases and controls. Our experience demonstrates that beyond PALB2, the prevalence of mutations in the remaining breast cancer predisposition genes is likely to be very low making definitive validation exceptionally challenging.

Keywords: predisposition, familial, exome sequencing, breast cancer

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1887 The Identification of Combined Genomic Expressions as a Diagnostic Factor for Oral Squamous Cell Carcinoma

Authors: Ki-Yeo Kim

Abstract:

Trends in genetics are transforming in order to identify differential coexpressions of correlated gene expression rather than the significant individual gene. Moreover, it is known that a combined biomarker pattern improves the discrimination of a specific cancer. The identification of the combined biomarker is also necessary for the early detection of invasive oral squamous cell carcinoma (OSCC). To identify the combined biomarker that could improve the discrimination of OSCC, we explored an appropriate number of genes in a combined gene set in order to attain the highest level of accuracy. After detecting a significant gene set, including the pre-defined number of genes, a combined expression was identified using the weights of genes in a gene set. We used the Principal Component Analysis (PCA) for the weight calculation. In this process, we used three public microarray datasets. One dataset was used for identifying the combined biomarker, and the other two datasets were used for validation. The discrimination accuracy was measured by the out-of-bag (OOB) error. There was no relation between the significance and the discrimination accuracy in each individual gene. The identified gene set included both significant and insignificant genes. One of the most significant gene sets in the classification of normal and OSCC included MMP1, SOCS3 and ACOX1. Furthermore, in the case of oral dysplasia and OSCC discrimination, two combined biomarkers were identified. The combined genomic expression achieved better performance in the discrimination of different conditions than in a single significant gene. Therefore, it could be expected that accurate diagnosis for cancer could be possible with a combined biomarker.

Keywords: oral squamous cell carcinoma, combined biomarker, microarray dataset, correlated genes

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1886 The Cleavage of DNA by the Anti-Tumor Drug Bleomycin at the Transcription Start Sites of Human Genes Using Genome-Wide Techniques

Authors: Vincent Murray

Abstract:

The glycopeptide bleomycin is used in the treatment of testicular cancer, Hodgkin's lymphoma, and squamous cell carcinoma. Bleomycin damages and cleaves DNA in human cells, and this is considered to be the main mode of action for bleomycin's anti-tumor activity. In particular, double-strand breaks are thought to be the main mechanism for the cellular toxicity of bleomycin. Using Illumina next-generation DNA sequencing techniques, the genome-wide sequence specificity of bleomycin-induced double-strand breaks was determined in human cells. The degree of bleomycin cleavage was also assessed at the transcription start sites (TSSs) of actively transcribed genes and compared with non-transcribed genes. It was observed that bleomycin preferentially cleaved at the TSSs of actively transcribed human genes. There was a correlation between the degree of this enhanced cleavage at TSSs and the level of transcriptional activity. Bleomycin cleavage is also affected by chromatin structure and at TSSs, the peaks of bleomycin cleavage were approximately 200 bp apart. This indicated that bleomycin was able to detect phased nucleosomes at the TSSs of actively transcribed human genes. The genome-wide cleavage pattern of the bleomycin analogues 6′-deoxy-BLM Z and zorbamycin was also investigated in human cells. As found for bleomycin, these bleomycin analogues also preferentially cleaved at the TSSs of actively transcribed human genes. The cytotoxicity (IC₅₀ values) of these bleomycin analogues was determined. It was found that the degree of enhanced cleavage at TSSs was inversely correlated with the IC₅₀ values of the bleomycin analogues. This suggested that the level of cleavage at the TSSs of actively transcribed human genes was important for the cytotoxicity of bleomycin and analogues. Hence this study provided a deeper understanding of the cellular processes involved in the cancer chemotherapeutic activity of bleomycin.

Keywords: anti-tumour activity, bleomycin analogues, chromatin structure, genome-wide study, Illumina DNA sequencing

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1885 Analysis of Resistance and Virulence Genes of Gram-Positive Bacteria Detected in Calf Colostrums

Authors: C. Miranda, S. Cunha, R. Soares, M. Maia, G. Igrejas, F. Silva, P. Poeta

Abstract:

The worldwide inappropriate use of antibiotics has increased the emergence of antimicrobial-resistant microorganisms isolated from animals, humans, food, and the environment. To combat this complex and multifaceted problem is essential to know the prevalence in livestock animals and possible ways of transmission among animals and between these and humans. Enterococci species, in particular E. faecalis and E. faecium, are the most common nosocomial bacteria, causing infections in animals and humans. Thus, the aim of this study was to characterize resistance and virulence factors genes among two enterococci species isolated from calf colostrums in Portuguese dairy farms. The 55 enterococci isolates (44 E. faecalis and 11 E. faecium) were tested for the presence of the resistance genes for the following antibiotics: erythromicyn (ermA, ermB, and ermC), tetracycline (tetL, tetM, tetK, and tetO), quinupristin/dalfopristin (vatD and vatE) and vancomycin (vanB). Of which, 25 isolates (15 E. faecalis and 10 E. faecium) were tested until now for 8 virulence factors genes (esp, ace, gelE, agg, cpd, cylA, cylB, and cylLL). The resistance and virulence genes were performed by PCR, using specific primers and conditions. Negative and positive controls were used in all PCR assays. All enterococci isolates showed resistance to erythromicyn and tetracycline through the presence of the genes: ermB (n=29, 53%), ermC (n=10, 18%), tetL (n=49, 89%), tetM (n=39, 71%) and tetK (n=33, 60%). Only two (4%) E. faecalis isolates showed the presence of tetO gene. No resistance genes for vancomycin were found. The virulence genes detected in both species were cpd (n=17, 68%), agg (n=16, 64%), ace (n=15, 60%), esp (n=13, 52%), gelE (n=13, 52%) and cylLL (n=8, 32%). In general, each isolate showed at least three virulence genes. In three E. faecalis isolates was not found virulence genes and only E. faecalis isolates showed virulence genes for cylA (n=4, 16%) and cylB (n=6, 24%). In conclusion, these colostrum samples that were consumed by calves demonstrated the presence of antibiotic-resistant enterococci harbored virulence genes. This genotypic characterization is crucial to control the antibiotic-resistant bacteria through the implementation of restricts measures safeguarding public health. Acknowledgements: This work was funded by the R&D Project CAREBIO2 (Comparative assessment of antimicrobial resistance in environmental biofilms through proteomics - towards innovative theragnostic biomarkers), with reference NORTE-01-0145-FEDER-030101 and PTDC/SAU-INF/30101/2017, financed by the European Regional Development Fund (ERDF) through the Northern Regional Operational Program (NORTE 2020) and the Foundation for Science and Technology (FCT). This work was supported by the Associate Laboratory for Green Chemistry - LAQV which is financed by national funds from FCT/MCTES (UIDB/50006/2020 and UIDP/50006/2020).

Keywords: antimicrobial resistance, calf, colostrums, enterococci

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1884 mRNA Biomarkers of Mechanical Asphyxia-Induced Death in Cardiac Tissue

Authors: Yan Zeng, Li Tao, Liujun Han, Tianye Zhang, Yongan Yu, Kaijun Ma, Long Chen

Abstract:

Mechanical asphyxia is one of the main cause of death; however, death by mechanical asphyxia may be difficult to prove in court, particularly in cases in which corpses exhibit no obvious signs of asphyxia. To identify a credible biomarker of asphyxia, we first examined the expression levels of all the mRNAs in human cardiac tissue specimens subjected to mechanical asphyxia and compared these expression levels with those of the corresponding mRNAs in specimens subjected to craniocerebral injury. A total of 119 differentially expressed mRNAs were selected and the expression levels of these mRNAs were examined in 44 human cardiac tissue specimens subjected to mechanical asphyxia, craniocerebral injury, hemorrhagic shock and other causes of death. We found that DUSP1 and KCNJ2 were up-regulated in tissue specimens of mechanical asphyxia compared with control tissues, with no significant correlation between age, environmental temperature and PMI, indicating that DUSP1 and KCNJ2 may associate with mechanical asphyxia-induced death and can thus serve as useful biomarkers of death by mechanical asphyxia.

Keywords: mechanical asphyxia, biomarkers, DUSP1, KCNJ2, cardiac tissue

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