Search results for: TP53 mutation/polymorphism

Authors: Helan Satish, M. Ramasubba Reddy

Abstract:

The simulation of an endocardial cell for gene mutation in the cardiac sodium ion channel NaV1.5, encoded by SCN5A gene, is discussed. The characterization of Brugada Syndrome by loss of function effect on SCN5A mutation due to L812Q mutant present in the DII-S4 transmembrane region of the NaV1.5 channel protein and its effect in an endocardial cell is studied. Ten Tusscher model of human ventricular action potential is modified to incorporate the changes contributed by L812Q mutant in the endocardial cells. Results show that BrS-associated SCN5A mutation causes reduction in the inward sodium current by modifications in the channel gating dynamics such as delayed activation, enhanced inactivation, and slowed recovery from inactivation in the endocardial cell. A decrease in the inward sodium current was also observed, which affects depolarization phase (Phase 0) that leads to reduction in the spike amplitude of the cardiac action potential.

Keywords: SCN5A gene mutation, sodium channel, Brugada syndrome, cardiac arrhythmia, action potential

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Authors: Ihtisham Bukhari

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In the current study, we enrolled a 46, XY phenotypically female patient bearing testes in her inguinal canal. DNA sequencing of the AR gene detected a missense mutation C.1715A > G (p. Y572C) in exon 2 which is already known to cause Complete androgen insensitivity syndrome (CAIS). We further studied the effects of this mutation on the testicular histopathology of the patient. No spermatocytes were seen in the surface spreading of testicular tissues while H&E staining showed that seminiferous tubules predominantly have only Sertoli cells. To confirm this meiotic failure is likely due to the current AR mutation we performed mRNA expression of genes associated with AR pathway, expression and location of the associated proteins in testicular tissues. Western blot and real-time PCR data showed that the patient had high levels of expression of AMH, SOX9, and INNB in testis. Tubules were stained with SOX9 and AMH which revealed Sertoli cell maturation arrest. Therefore, we suggest that AR mutation enhances AMH expression which ultimately leads to failure in the maturation of Sertoli cells and failure in spermatogenesis.

Keywords: androgen receptor, spermatogenesis, infertility, Sertoli cell only syndrome

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Authors: M. Reni Sagayaraj, S. Anand Gnana Selvam, R. Reynald Susainathan

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We analyze stochastic integrals associated with a mutation process. To be specific, we describe the cell population process and derive the differential equations for the joint generating functions for the number of mutants and their integrals in generating functions and their applications. We obtain first-order moments of the processes of the two-way mutation process in first-order moment structure of X (t) and Y (t) and the second-order moments of a one-way mutation process. In this paper, we obtain the limiting behaviour of the integrals in limiting distributions of X (t) and Y (t).

Keywords: stochastic integrals, single–server queue model, catastrophes, busy period

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Authors: Shahram Nanekarania, Majid Goodarzia

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Calpastatin and callipyge have been known as one of the candidate genes in meat quality and quantity. Calpastatin gene has been located to chromosome 5 of sheep and callipyge gene has been localized in the telomeric region on ovine chromosome 18. The objective of this study was identification of calpastatin and callipyge genes polymorphism and analysis of genotype structure in population of Lori sheep kept in Iran. Blood samples were taken from 120 Lori sheep breed and genomic DNA was extracted by salting out method. Polymorphism was identified using the PCR-RFLP technique. The PCR products were digested with MspI and FaqI restriction enzymes for calpastatin gene and callipyge gene, respectively. In this population, three patterns were observed and AA, AB, BB genotype have been identified with the 0.32, 0.63, 0.05 frequencies for calpastatin gene. The results obtained for the callipyge gene revealed that only the wild-type allele A was observed, indicating that only genotype AA was present in the population under consideration.

Keywords: polymorphism, calpastatin, callipyge, PCR-RFLP, Lori sheep

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Authors: Arab M., Ait Abdallah M., Zeraoulia N., Boumaza H., Aoutia M., Griene L., Ait Abdelkader B.,

Abstract:

breast and ovarian cancer are respectively the first and fourth leading causes of cancer among women in Algeria. A family story of cancer in the most important risk factor, and in most cases of families with breast and /or ovarian cancer, the pattern of cancer family can be attributed to mutation in BRCA1/2genes. objectibes: the aim of our study in to investigate the spectrum of BRCA1/2 germiline mutation in familial breast and /or ovarian cancer and to determine the prevalence and the nature of BRCA1/2mutation in Algeria methods: we deremined the prevalence of BRCA1/2 mutation within a cohort of 161 probands selected according the eisinger score double stranded sanger sequencing of all coding exons of BRCA1/2including flanking intronic region were performed results: we identified a total of 23 distinct deleterious mutations (class5) 12 differents mutations in BRCA1(52%) and 11 in BRCA2(48%). 78% (18/23) were protein truncating and 22%(5/23) missens mutations.3 novel deleterious mutations have been identified, which have not been described in public mutation database. one new mutation were found in two unrelated patients. the overall mutation detection rate in our study is 28,5%(46/161).more over, an UVS c7783 located in BRCA2 is found in two unrelated probands and segregate in the 02 families/ conclusion: our results sugget of large spectrum of BRCA1/2 mutation in Algerian breast/ovarian cancer family. The nature and prevalence of BRCA1/2mutation in algerian families are ongoing in a larger study, 80 probands are to day under investigation. This study which may therefore identify the genetic particularity of Algerian breast /ovarian cancer.

Keywords: BRCA1/2 mutations, hereditary breast cancer, algerian women, prvalence

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Authors: Mehrdad Sheikhvatan, Mohammad Ali Boroumand, Mehrdad Behmanesh, Shayan Ziaee

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Contradictory results have been obtained regarding the role of integrin, beta 3 (ITGB3) gene polymorphisms in occurrence of acute myocardial infarction (MI) in patients with coronary artery disease (CAD). Hence, we aimed to assess the association between 1565C/T polymorphism of ITGB3 gene and increased risk for acute MI in patients who suffered premature CAD in Iranian population. Our prospective study included 1000 patients (492 men and 508 women aged 21 to 55 years) referred to Tehran Heart center during a period of four years from 2008 to 2011 with the final diagnosis of premature CAD and classified into two groups with history of MI (n = 461) and without of MI (n = 539). The polymorphism variants were determined by PCR-RFLP technique by entering 10% of randomized samples and then genotyping of the polymorphism was also conducted by High Resolution Melting (HRM) method. Among study samples, 640 were followed with a median follow-up time 45.74 months for determining association of long-term major adverse cardiac events (MACE) and genotypes of polymorphisms. There was no significant difference in the frequency of 1565C/T polymorphism between the MI and non-MI groups. The frequency of wild genotype was 69.2% and 72.2%, the frequency of homozygous genotype was 21.3% and 18.4%, and the frequency of mutant genotype was 9.5% and 9.5%, respectively (p=0.505). Results were also similar when adjusted for covariates in a multivariate logistic regression model. No significant difference was also found in total-MACE free survival rate between the patients with different genotypes of 1565C/T polymorphism in both MI and non-MI group. The carriage of the 1565C/T polymorphism of ITGB3 gene seems unlikely to be a significant risk factor for the development of MI in Iranian patients with premature CAD. The presence of this ITGB3 gene polymorphism may not also predict long-term cardiac events.

Keywords: coronary artery disease, myocardial infarction, gene, integrin, beta 3, polymorphism

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Authors: Omar Youssef, Aija Knuuttila, Paivi Piirilä, Virinder Sarhadi, Sakari Knuutila

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Exhaled breath condensate (EBC) is a unique sample that allows studying different genetic changes in lung carcinoma through a non-invasive way. With the aid of next generation sequencing (NGS) technology, analysis of genetic mutations has been more efficient with increased sensitivity for detection of genetic variants. In order to investigate the possibility of applying this method for cancer diagnostics, mutations in EBC DNA from lung cancer patients and healthy individuals were studied by using NGS. The key aim is to assess the feasibility of using this approach to detect clinically important mutations in EBC. EBC was collected from 20 healthy individuals and 9 lung cancer patients (four lung adenocarcinomas, four 8 squamous cell carcinoma, and one case of mesothelioma). Mutations in hotpot regions of 22 genes were studied by using Ampliseq Colon and Lung cancer panel and sequenced on Ion PGM. Results demonstrated that all nine patients showed a total of 19 cosmic mutations in APC, BRAF, EGFR, ERBB4, FBXW7, FGFR1, KRAS, MAP2K1, NRAS, PIK3CA, PTEN, RET, SMAD4, and TP53. In controls, 15 individuals showed 35 cosmic mutations in BRAF, CTNNB1, DDR2, EGFR, ERBB2, FBXW7, FGFR3, KRAS, MET, NOTCH1, NRAS, PIK3CA, PTEN, SMAD4, and TP53. Additionally, 45 novel mutations not reported previously were also seen in patients’ samples, and 106 novel mutations were seen in controls’ specimens. KRAS exon 2 mutations G12D was identified in one control specimen with mutant allele fraction of 6.8%, while KRAS G13D mutation seen in one patient sample showed mutant allele fraction of 17%. These findings illustrate that hotspot mutations are present in DNA from EBC of both cancer patients and healthy controls. As some of the cosmic mutations were seen in controls too, no firm conclusion can be drawn on the clinical importance of cosmic mutations in patients. Mutations reported in controls could represent early neoplastic changes or normal homeostatic process of apoptosis occurring in lung tissue to get rid of mutant cells. At the same time, mutations detected in patients might represent a non-invasive easily accessible way for early cancer detection. Follow up of individuals with important cancer mutations is necessary to clarify the significance of these mutations in both healthy individuals and cancer patients.

Keywords: exhaled breath condensate, lung cancer, mutations, next generation sequencing

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Authors: Nermin Gozukirmizi, Buket Cakmak, Sevgi Marakli

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Sireviruses are genera of copia LTR retrotransposons with a unique genome structure among retrotransposons. Barley (Hordeum vulgare L.) is an economically important plant and has been studied as a model plant regarding its short annual life cycle and seven chromosome pairs. In this study, we used mature barley embryos, 10-day-old roots and 10-day-old leaves derived from the same barley plant to investigate SIRE1 retrotransposon movements by Inter-Retrotransposon Amplified Polymorphism (IRAP) technique. We found polymorphism rates between 0-64% among embryos, roots and leaves. Polymorphism rates were detected to be 0-27% among embryos, 8-60% among roots, and 11-50% among leaves. Polymorphisms were observed not only among the parts of different individuals, but also on the parts of the same plant (23-64%). The internal domains of SIRE1 (gag, env and rt) were also analyzed in the embryos, roots and leaves. Analysis of band profiles showed no polymorphism for gag, however, different band patterns were observed among samples for rt and env. The sequencing of SIRE1 gag, env and rt domains revealed 79% similarity for gag, 95% for env and 84% for rt to Ty1-copia retrotransposons. SIRE1 retrotransposon was identified in the soybean genome and has been studied on other plants (maize, rice, tomatoe etc.). This study is the first detailed investigation of SIRE1 in barley genome. The obtained findings are expected to contribute to the comprehension of SIRE1 retrotransposon and its role in barley genome.

Keywords: barley, polymorphism, retrotransposon, SIRE1 virus

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Authors: Claudia Patricia Beltran Miranda, Mónica López Anaya, Mónica Navarro Meza, Maria Leonor Valderrama Chairez

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Background: Mexico occupies the second world-wide place of morbid obese people (10- 12 million). Factors that predispose the development of MO are genetic, environmental, physiological, psycho-social and behavioral (nourishing habits). Objective: Associate Gln223Arg polymorphism of gene LEPR, levels of leptina and nourishing habits with the presence of morbid obesity in adolescents of the south of Jalisco (México). Methods: In 41 adolescents (18 normal weight and 23 morbid obesity) of 12 to 19 years of age, both sexes that were measure size and weight with tanita scale and stadimeter to determine IMC. Morbid obesity was determined by tables of the WHO and was established with a standard deviation >3. The Gln223Arg polymorphism have been identify by PCR and leptina levels by ELISA. Nourishing habits were evaluate by the questionnaire the Adolescent Food Habits Checklist. The statistical analysis was performed to compare mean scores obtained from the questionnaire when we compare morbid obesity vs. normal weight adolescents with p=0.03 and a significance of 95%. Results: frequencies alellics and genics were not stadistics significatives p= 0,011 and p=0,279 respectly when were compared between normal weight adolescents and morbib obesity Leptin levels and nourishing habits were associated with morbid obesity. The polymorphism not shown significance with morbid obesity. Conclusions: Dietary habits and leptin levels in adolescents are important factor that predisposes the development of obesity in adolescents. The presence of the polymorphism are not associated with morbid obesity in these subjects.

Keywords: leptin, nourishing habits, morbid obesity, polymorphism

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Authors: Małgorzata Wrzosek, Nina Baruch, Beata Jabłonowska-Lietz

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Background: The fat mass & obesity-associated (FTO) gene is linked to an increased risk of obesity. However, the relation between rs9939609 and eating behaviors or energy expenditure is not fully elucidated. The aim of this study was to investigate the relationship between the rs9939609 polymorphism of the FTO gene and the postprandial satiety, sweets intake, physical activity and depressive symptoms in patients with obesity. Methods: The study group consisted of 585 subjects with a BMI of 42.97.0 kg/m². The rs9939609 polymorphism of the FTO gene was examined using real time – PCR method. The severity of depressive symptoms was assessed with the Beck Depression Inventory (BDI-II). Information was obtained about demographics, eating habits and lifestyle. Results: More than half (63.5%) of the patients reported consumption of sweets between main meals and 30% declared high and very high postprandial satiety and the frequency of TA/AA carriers in rs9939609 (FTO) compared with TT carriers was similar. Significantly lower BDI-II scores were found in subjects with higher level of physical activity and it was seen amongst patients with the AA and AT genotypes of the FTO rs9939609 polymorphism. Conclusion: Obesity is a highly heritable trait, but eating habits also appear as major factors affecting obesity development.

Keywords: FTO polymorphism, physical activity, obesity, depression, postprandial satiety, sugary foods, sweets

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Authors: Roya Bakhtiar, Alireza Abdolmohammadi, Hadi Hajarian, Zahra Nikousefat, Davood, Kalantar-Neyestanaki

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The objective of the present study was to determine the polymorphism in the leptin (332G>A) and its association with biometric traits in Sanjabi sheep. For this purpose, blood samples from 96 rams were taken, and tail length, width tail, circumference tail, body length, body width, and height were simultaneously recorded. PCR was performed using specific primer to amplify 463 bp fragment including exon 3 of leptin gene, and PCR products were digested by Cail restriction enzymes. The 332G>A (at 332th nucleotide of exon 3 leptin gene) that caused an amino acid change from Arg to Gln was detected by Cail (CAGNNNCTG) endonuclease, as the endonuclease cannot cut this region if G nucleotide is located in this position. Three genotypes including GG (463), GA (463, 360and 103 bp) and GG (360 bp and 103 bp) were identified after digestion by enzyme. The estimated frequencies of three genotypes including GG, GA, and AA for 332G>A locus were 0.68, 0.29 and 0.03 and those were 0.18 and 0.82 for A and G alleles, respectively. In the current study, chi-square test indicated that 332G>A positions did not deviate from the Hardy–Weinberg (HW) equilibrium. The most important reason to show HW equation was that samples used in this study belong to three large local herds with a traditional breeding system having random mating and without selection. Shannon index amount was calculated which represent an average genetic variation in Sanjabi rams. Also, heterozygosity estimated by Nei index indicated that genetic diversity of mutation in the leptin gene is moderate. Leptin gene polymorphism in the 332G>A had significant effect on body length (P<0.05) trait, and individuals with GA genotype had significantly the higher body length compared to other individuals. Although animals with GA genotype had higher body width, this difference was not statistically significant (P>0.05). This non-synonymous SNP resulted in different amino acid changes at codon positions111(R/Q). As leptin activity is localized, at least in part, in domains between amino acid residues 106-1406, it is speculated that the detected SNP at position 332 may affect the activity of leptin and may lead to different biological functions. Based to our results, due to significant effect of leptin gene polymorphism on body size traits, this gene may be used a candidate gene for improving these traits.

Keywords: body size, Leptin gene, PCR-RFLP, Sanjabi sheep

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Authors: Hossein Rassi, Marjan Moradi Fard, Masoud Houshmand

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Background: Cervical cancer is multistep disease that is thought to result from an interaction between genetic background and environmental factors. Human papillomavirus (HPV) infection is the leading risk factor for cervical intraepithelial neoplasia (CIN) and cervical cancer. In other hand, some of p53 and miRNA polymorphism may plays an important role in carcinogenesis. This study attempts to clarify the relation of p53 genotypes and miR-146a rs2910164 polymorphism in cervical lesions. Method: Forty two archival samples with cervical lesion retired from Khatam hospital and 40 sample from healthy persons used as control group. A simple and rapid method was used to detect the simultaneous amplification of the HPV consensus L1 region and HPV-16,-18, -11, -31, 33 and -35 along with the b-globin gene as an internal control. We use Multiplex PCR for detection of P53 and miR-146a rs2910164 genotypes in our lab. Finally, data analysis was performed using the 7 version of the Epi Info(TM) 2012 software and test chi-square(x2) for trend. Results: Cervix lesions were collected from 42 patients with Squamous metaplasia, cervical intraepithelial neoplasia, and cervical carcinoma. Successful DNA extraction was assessed by PCR amplification of b-actin gene (99bp). According to the results, p53 GG genotype and miR-146a rs2910164 CC genotype was significantly associated with increased risk of cervical lesions in the study population. In this study, we detected 13 HPV 18 from 42 cervical cancer. Conclusion: The connection between several SNP polymorphism and human virus papilloma in rare researches were seen. The reason of these differences in researches' findings can result in different kinds of races and geographic situations and also differences in life grooves in every region. The present study provided preliminary evidence that a p53 GG genotype and miR-146a rs2910164 CC genotype may effect cervical cancer risk in the study population, interacting synergistically with HPV 18 genotype. Our results demonstrate that the testing of p53 codon 72 polymorphism genotypes and miR-146a rs2910164 polymorphism genotypes in combination with HPV18 can serve as major risk factors in the early identification of cervical cancers. Furthermore, the results indicate the possibility of primary prevention of cervical cancer by vaccination against HPV18 in Iran.

Keywords: cervical cancer, HPV18, p53 codon 72 polymorphism, miR-146a rs2910164 polymorphism

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Authors: Mahya Naghipoor

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Purpose: Non-small cell lung cancer (NSCLC) is the most common lung cancer type. Epidermal growth factor receptor (EGFR) mutation is the main reason which causes NSCLC. Computed tomography (CT) is used for diagnosis and prognosis of lung cancers because of low price and little invasion. Semantic analyses of qualitative CT features are based on visual evaluation by radiologist. However, the naked eye ability may not assess all image features. On the other hand, radiomics provides the opportunity of quantitative analyses for CT images features. The aim of this review study was comparing accuracy of semantic and radiomics features in prognosis of EGFR mutation in NSCLC. Methods: For this purpose, the keywords including: non-small cell lung cancer, epidermal growth factor receptor mutation, semantic, radiomics, feature, receiver operating characteristics curve (ROC) and area under curve (AUC) were searched in PubMed and Google Scholar. Totally 29 papers were reviewed and the AUC of ROC analyses for semantic and radiomics features were compared. Results: The results showed that the reported AUC amounts for semantic features (ground glass opacity, shape, margins, lesion density and presence or absence of air bronchogram, emphysema and pleural effusion) were %41-%79. For radiomics features (kurtosis, skewness, entropy, texture, standard deviation (SD) and wavelet) the AUC values were found %50-%86. Conclusions: In conclusion, the accuracy of radiomics analysis is a little higher than semantic in prognosis of EGFR mutation in NSCLC.

Keywords: lung cancer, radiomics, computer tomography, mutation

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Authors: Mohammed N. Al Kindi, Mazin Al Khabouri, Khalsa Al Lamki, Tommasso Pappuci, Giovani Romeo, Nadia Al Wardy

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Hereditary hearing loss is a heterogeneous group of complex disorders with an overall incidence of one in every five hundred newborns presented as syndromic and non-syndromic forms. Cadherin-related 23 (CDH23) is one of the listed deafness causative genes. CDH23 is found to be expressed in the stereocilia of hair cells and the retina photoreceptor cells. Defective CDH23 has been associated mostly with prelingual severe-to-profound sensorineural hearing loss (SNHL) in either syndromic (USH1D) or non-syndromic SNHL (DFNB12). An Omani family diagnosed clinically with severe-profound sensorineural hearing loss was genetically analysed by whole exome sequencing technique. A novel homozygous missense variant, c.A7451C (p.D2484A), in exon 53 of CDH23 was detected. One hundred and thirty control samples were analysed where all were negative for the detected variant. The variant was analysed in silico for pathogenicity verification using several mutation prediction software. The variant proved to be a pathogenic mutation and is reported for the first time in Oman and worldwide. It is concluded that in silico mutation prediction analysis might be used as a useful molecular diagnostics tool benefiting both genetic counseling and mutation verification. The aspartic acid 2484 alanine missense substitution might be the main disease-causing mutation that damages CDH23 function and could be used as a genetic hearing loss marker for this particular Omani family.

Keywords: Cdh23, d2484a, in silico, Oman

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Authors: Apichai Sawisit, Sirima Suvarnakuta Jantama, Sunthorn Kanchanatawee, Lonnie O. Ingram, Kaemwich Jantama

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Escherichia coli KJ122 was engineered to produce succinate from glucose using the wild type GalP for glucose uptake instead of the native phosphotransferase system (ptsI mutation). This strain ferments 10% (w/v) xylose poorly. Mutants were selected by serial transfers in AM1 mineral salts medium with 10% (w/v) xylose. Evolved mutants exhibited a similar improvement, co-fermentation of an equal mixture of xylose and glucose. One of these, AS1600a, produced 84.26±1.37 g/L succinate, equivalent to that produced by the parent (KJ122) strain from 10% glucose (85.46±1.78 g/L). AS1600a was sequenced and found to contain a mutation in galactose permease (GalP, G236D). Expressing the galP* mutation gene in KJ122ΔgalP resembled the xylose utilization phenotype of the mutant AS1600a. The strain AS1600a and KJ122ΔgalP (pLOI5746; galP*) also co-fermented a mixture of glucose, xylose, arabinose, and galactose in sugarcane bagasse hydrolysate for succinate production.

Keywords: xylose, furfural, succinat, sugarcane bagasse, E. coli

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Authors: Sanjeev Kumar Shukla, Govind Singh, Prabhat Pant Shahzad Ahmad

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Background: Graves’ disease is an autoimmune disorder with a genetic predisposition, and CD40 plays a pathogenic role in various autoimmune diseases. A single nucleotide polymorphism at position –1 of the Kozak sequence of the 5 untranslated regions of the CD40 gene of exon 1 has been reported to be associated with the development of Graves’ Disease. Objective: The aim of the present study was to investigate whether CD40 gene polymorphism confers susceptibility to Graves’ disease in the Kumaon region. CD40 gene polymorphisms were studied in Graves’ Disease patients (n=50) and healthy control subjects without anti-thyroid autoantibodies or a family history of autoimmune disorders (n=50). Material and Method: CD40 gene polymorphisms were studied in fifty Graves’ Disease patients and fifty healthy control subjects. All samples were collected from STG Hospital, Haldwani, Nainital. A C/T polymorphism at position –1 of the CD40 gene was measured using the polymerase chain reaction-restriction fragment length polymorphism. Results: There was no significant difference in allele or genotype frequency of the CD40 SNP between Graves’ Disease and control subjects. There was a significant decrease in the TT genotype frequency in the Graves’ Disease patients who developed Graves’ Disease after 40 years old than those under 40 years of age. These data suggest that the SNP of the CD40 gene is associated with susceptibility to the later onset of Graves’ Disease. Conclusion: The CD40 gene was a different susceptibility gene for Graves’ Disease within certain families because it was both linked and associated with Graves’ Disease.

Keywords: autoimmune diseases, pathogenesis, diagnosis, therapy

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Authors: Rouhollah Radjabi, Mojtaba Zarei, Elham Sanatgar, Hossein Shouhani

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RAPD molecular markers in order to discrimination of the Iranian native Bombyx mori silkworm breeds were used. DNA extraction using phenol - chloroform was and the qualitative and quantitative measurements of extracted DNA and its dilution, the obtained bands on agarose gel 1.5 percent were marked and analyzed. Results showed that the bands are observed between 250-2500 bp and most bands have been observed as Gilani-orange, the lowest bands observed are Khorasani-lemon. Primer 3 with 100% polymorphism with the highest polymorphism and primer 2 with 61.5 polymorphism had the lowest percentage of polymorphism. Cluster analysis of races and placed them in three main groups, races Gilani - orange, Baghdad and Khorasani -pink if the first group, camel's thorn, Herati - yellow race was alone in the second group and Khorasani – lemon was alone in the third group. The greatest similarity between the races, between Khorasani- pink and Baghdad (0.64). RAPD markers have been determined different silkworm races based on various morphological or economic characteristics except geographic origin.

Keywords: silkworm, molecular marker, RAPD, Iran

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Authors: Ogbonnaya O. Iroanya, Samson T. Fakorede, Osamudiamen J. Edosa, Hadiat A. Azeez

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The human mitochondrial DNA (mtDNA) is about 17 kbp circular DNA fragments found within the mitochondria together with smaller fragments of 1200 bp known as the control region. Knowledge of variation within populations has been employed in forensic and molecular anthropology studies. The study was aimed at investigating the polymorphic nature of the two hypervariable segments (HVS) of the mtDNA, i.e., HVS1 and HVS2, and to determine the haplogroup distribution among individuals resident in Lagos, Southwestern Nigeria. Peripheral blood samples were obtained from sixty individuals who are not related maternally, followed by DNA extraction and amplification of the extracted DNA using primers specific for the regions under investigation. DNA amplicons were sequenced, and sequenced data were aligned and compared to the revised Cambridge Reference Sequence (rCRS) GenBank Accession number: NC_012920.1) using BioEdit software. Results obtained showed 61 and 52 polymorphic nucleotide positions for HVS1 and HVS2, respectively. While a total of three indels mutation were recorded for HVS1, there were seven for HVS2. Also, transition mutations predominate nucleotide change observed in the study. Genetic diversity (GD) values for HVS1 and HVS2 were estimated to be 84.21 and 90.4%, respectively, while random match probability was 0.17% for HVS1 and 0.89% for HVS2. The study also revealed mixed haplogroups specific to the African (L1-L3) and the Eurasians (U and H) lineages. New polymorphic sites obtained from the study are promising for human identification purposes.

Keywords: hypervariable region, indels, mitochondrial DNA, polymorphism, random match probability

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Authors: Hossein Rassi, Marjan Moradi fard, Masoud Houshmand

Abstract:

Background: Cervical cancer is multistep disease that is thought to result from an interaction between genetic background and environmental factors. Human Papillomavirus (HPV) infection is the leading risk factor for Cervical Intraepithelial Neoplasia (CIN) and cervical cancer. In other hand, some of p53 and miRNA polymorphism may plays an important role in carcinogenesis. This study attempts to clarify the relation of p53 genotypes and miR-146a rs2910164 polymorphism in cervical lesions. Method: Forty two archival samples with cervical lesion retired from Khatam hospital and 40 sample from healthy persons used as control group. A simple and rapid method was used to detect the simultaneous amplification of the HPV consensus L1 region and HPV-16,-18, -11, -31, 33, and -35 along with the b-globin gene as an internal control. We use Multiplex PCR for detection of P53 and miR-146a rs2910164 genotypes in our lab. Finally, data analysis was performed using the 7 version of the Epi Info(TM) 2012 software and test chi-square(x2) for trend. Results: Cervix lesions were collected from 42 patients with Squamous metaplasia, cervical intraepithelial neoplasia, and cervical carcinoma. Successful DNA extraction was assessed by PCR amplification of b-actin gene (99 bp). According to the results, p53 GG genotype and miR-146a rs2910164 CC genotype was significantly associated with increased risk of cervical lesions in the study population. In this study, we detected 13 HPV 18 from 42 cervical cancer. Conclusion: The connection between several SNP polymorphism and human virus papilloma in rare researches were seen. The reason of these differences in researches' findings can result in different kinds of races and geographic situations and also differences in life grooves in every region. The present study provided preliminary evidence that a p53 GG genotype and miR-146a rs2910164 CC genotype may effect cervical cancer risk in the study population, interacting synergistically with HPV 18 genotype. Our results demonstrate that the testing of p53 codon 72 polymorphism genotypes and miR-146a rs2910164 polymorphism genotypes in combination with HPV18 can serve as major risk factors in the early identification of cervical cancers. Furthermore, the results indicate the possibility of primary prevention of cervical cancer by vaccination against HPV18 in Iran.

Keywords: cervical cancer, miR-146a rs2910164 polymorphism, p53 polymorphism, intraepithelial, neoplasia, HPV

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Authors: Manaphol Kulpraneet, Anirut Limtrakul, Surangrat Srisurapanon, Piyatida Tangteerawatana

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Tuberculosis (TB) remains a global health care disease world-wide. Control of the global TB epidemic has been impaired by the lack of an effective vaccine, by the emergence of drug resistant forms of Mycobacterium tuberculosis and by lack of sensitive and rapid diagnostics. Cytokines play a major role in defense against M. tuberculosis infection. Polymorphisms in the genes encoding various cytokines have been associated with tuberculosis susceptibility. Polymorphisms of the regulatory cytokine gene, the interleukin (IL)-10 is associated with the risk of tuberculosis (TB) in different populations. However, IL-10 gene polymorphism and susceptibility to TB in Thai is still unknown. The purpose of this study was to evaluate whether the common IL-10 promoter gene polymorphisms are associated with TB in Thai population. Forty eight patients with newly diagnosed pulmonary tuberculosis were studied. DNA samples were extracted from leukocytes and used to investigate -1087A/G, -819C/T, -252C/A (rs1800896, rs1800871, rs1800872) in IL-10 gene using restriction fragment length polymorphism (PCR-RFLP) methods. In this study, the genotype and allele frequencies of IL-10-1087A/G, -819C/T, -252C/A polymorphism did not significantly different between TB patients and healthy controls ((genotype: p=0.38, p=0.92, p=1; allele: p=0.57, p=0.77, p=0.89, respectively). The lack of association between common IL-10 promoter polymorphisms and TB susceptibility in this study may provide clue for better understanding of IL-10-1087A/G, -819C/T, -252C/A polymorphism and TB susceptibility in Thai population, which might facilitate the rationale design of vaccines. However, further studies in large scales population are required for confirmation.

Keywords: IL-10, cytokines, single nucleotide polymorphism (SNP), tuberculosis

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Authors: Rosetta Moshirian Farahi, Ahya Abdi Ali, Sara Gharavi

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The major mechanism of Pseudomonas aeruginosa resistance to fluoroquinolones is the alteration of target enzymes, type II and IV topoisomerases due to mutations in the quinolone resistance-determining regions (QRDR) of the gyrA and parC genes coding A subunits of these enzymes. 37 isolates from patients with burn wounds and 20 isolates from blood, urine and sputum specimen were selected to evaluate mutations involved in antibiotic resistance and were subsequently verified for their resistance to ciprofloxacin. QRDRs regions of gyrA and parC were amplified by polymerase chain reaction (PCR) and were subsequently sequenced. 90% of isolates with MIC≥8 µg/ml to ciprofloxacin had a mutation in gyrA gene in which threonine at position 83 changed to isoleucine. 87.5% of isolates had mutation in parC, Serine 87 changed. 75% had Ser87Leu and 12.5% possessed Serin87Trp. Various silent mutations were also detected such as Val103Val, Ala118Ala, Ala136Ala, His132His in gyrA and Ala115Ala in parC. The data indicates that the common mutation in gyrA is Thr83Ile and in parC is Ser87Leu/Trp. No individual parC mutation was observed while mutations in gyrA and parC occurred simultaneously and appears to be the main reason of high-level resistance to fluoroquinolones in patients with burn wounds and urine infection. The vast majority of P.aeruginosa isolates had mutation in parC which can play a crucial role in increased resistance of these isolates. This is a report of parC mutations from resistant P. aeruginosa isolates from Iran, Tehran.

Keywords: P. aeruginosa, fluoroquinolones, gyrA, parC, antibiotic resistance

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Authors: Huong M. T. Nguyen, Hoa A. P. Nguyen, Mai P. T. Nguyen, Ngoc D. Ngo, Van T. Ta, Hai T. Le, Chi V. Phan

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Wilson’s disease (WD) is an autosomal recessive disorder of the copper metabolism, which is caused by a mutation in the copper-transporting P-type ATPase (ATP7B). The mechanism of this disease is the failure of hepatic excretion of copper to bile, and leads to copper deposits in the liver and other organs. The ATP7B gene is located on the long arm of chromosome 13 (13q14.3). This study aimed to investigate the gene mutation in the Vietnamese patients with WD, and make a presymptomatic diagnosis for their familial members. Forty-three WD patients and their 65 siblings were identified as having ATP7B gene mutations. Genomic DNA was extracted from peripheral blood samples; 21 exons and exon-intron boundaries of the ATP7B gene were analyzed by direct sequencing. We recognized four mutations ([R723=; H724Tfs*34], V1042Cfs*79, D1027H, and IVS6+3A>G) in the sum of 20 detectable mutations, accounting for 87.2% of the total. Mutation S105* was determined to have a high rate (32.6%) in this study. The hotspot regions of ATP7B were found at exons 2, 16, and 8, and intron 14, in 39.6 %, 11.6 %, 9.3%, and 7 % of patients, respectively. Among nine homozygote/compound heterozygote siblings of the patients with WD, three individuals were determined as asymptomatic by screening mutations of the probands. They would begin treatment after diagnosis. In conclusion, 20 different mutations were detected in 43 WD patients. Of this number, four novel mutations were explored, including [R723=; H724Tfs*34], V1042Cfs*79, D1027H, and IVS6+3A>G. The mutation S105* is the most prevalent and has been considered as a biomarker that can be used in a rapid detection assay for diagnosis of WD patients. Exons 2, 8, and 16, and intron 14 should be screened initially for WD patients in Vietnam. Based on risk profile for WD, genetic testing for presymptomatic patients is also useful in diagnosis and treatment.

Keywords: ATP7B gene, mutation detection, presymptomatic diagnosis, Vietnamese Wilson’s disease

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Authors: Dedy Pratama, Akhmadu Muradi, Hilman Ibrahim, Patrianef Darwis, Alexander Jayadi Utama, Raden Suhartono, D. Suryandari, Luluk Yunaini, Tom Ch Adriani

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Objective: Diabetic Foot Ulcer (DFU) is one of the complications of Type 2 Diabetes Mellitus (T2DM) that can lead to disability and death. Inadequate vascularization condition will affect healing process of DFU. Therefore, we investigated the expression of polymorphism TGF- β1 in the relation of the occurrence of DFU in T2DM. Methods: We designed a case-control study to investigate the polymorphism TGF- β1 gene 1800469 C > T and 1982073 C > T in T2DM in Cipto Mangunkusumo National Hospital (RSCM) Jakarta from June to December 2016. We used PCR techniques and compared the results in a group of T2DM patients with DFU as the case study and without DFU as the control group. Results: There were 203 patients, 102 patients with DFU and 101 patients control without DFU. 49,8% is male and 50,2% female with mean age about 56 years. Distribution of wild-type genotype TGF-B1 1800469 C > T wild type CC was found in 44,8%, the number of mutant heterozygote CT was 10,8% and mutant homozygote is 11,3%. Distribution of TGF-B1 1982073 C>T wild type CC was 32,5%, mutant heterozygote is 38,9% and mutant homozygote 25,1%. Conclusion: Distribution of alleles from TGF-B1 1800469 C > T is C 75% and T 25% and from TGF-B1 1982073 C > T is C53,8% and T 46,2%. In the other word polymorphism TGF- β1 plays a role in the occurrence and healing process of the DFU in T2DM patients.

Keywords: diabetic foot ulcers, diabetes mellitus, polymorphism, TGF-β1

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Authors: Seraj Makkawi, Abdulaziz A. Alqarni, Himyan Alghaythee, Suzan Y. Alharbi, Anmar Fatani, Reem Adas, Ahmad R. Abuzinadah

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Amyotrophic lateral sclerosis (ALS), also known as Lou Gehrig's disease, is a neurodegenerative disease that involves both the upper and lower motor neurons. Familial ALS, including superoxide dismutase 1 (SOD1) mutation, accounts for 5-10% of all cases of ALS. Typically, the symptoms of ALS are purely motor, though coexistent sensory symptoms have been reported in rare cases. In this report, we describe the case of a 47- year-old man who presented with progressive bilateral lower limb weakness and numbness for the last four years. A nerve conduction study (NCS) showed evidence of coexistent axonal sensorimotor polyneuropathy in addition to the typical findings of ALS in needle electromyography. Genetic testing confirmed the diagnosis of familial ALS secondary to the SOD1 genetic mutation. This report highlights that the presence of sensory symptoms should not exclude the possibility of ALS in an appropriate clinical setting.

Keywords: Saudi Arabia, polyneuropathy, SOD1 gene mutation, familial amyotrophic lateral sclerosis, amyotrophic lateral sclerosis

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Authors: Shahram Nanekarani, Majid Goodarzi, Majid Khosravi

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The present study aimed at analyzing of ovine major histocompatibility complex class II (Ovar II) DRB1 gene second exon in Lori Sheep breed. The MHC plays a central role in the control of disease resistance and immunological response. Genomic DNA from blood samples of 124 sheep was extracted and a 296 bp MHC exon 2 fragment was amplified using polymerase chain reaction. PCR products were characterized by the restriction fragment length polymorphism technique using Hin1I restriction enzyme. The PCRRFLP patterns showed three genotypes, AA, AB and BB with frequency of 0.282, 0.573 and 0.145, respectively. There was no significant (P > 0.05) deviation from Hardy–Weinberg equilibrium for this locus in this population. The results of the present study indicate that exon 2 of the Ovar-DRB1 gene is highly polymorphic in Lori sheep and could be considered as an important marker assisted selection, for improvement of immunity in sheep.

Keywords: MHC-DRB1 gene, polymorphism, PCR-RFLP, lori sheep

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Authors: R. Frikha, H. Kamoun

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Background:Chronicmyeloidleukemia (CML), a hematologicaldisease, ischaracterized by t (9; 22) and relatedoncogene BCR-ABL formation. Although Tyrosine kinase inhibitors (TKIs) have revolutionized the treatment of CML, resistanceoccurs and possibly médiates by mutation in severalgenesindependently of the bcr-abl1 kinase mechanism. it has been reportedthat JAK2V617F/BCR-ABL double positivitymaybe a potential marker of resistance in CML. Aims: This studywasinvestigated the JAK2V617F mutation in TKI-resistant CML patients. Methods: A retrospectivestudywasconducted in the Hospital University of Sfax, south of Tunisia, including all CML TKI-resistant patients. A Sanger sequencingwasperformedusing a high-fidelity DNA polymerase. Results:Nineresistant CP-CML patients wereenrolled in thisstudy. The JAK2V617F mutation wasdetectedin 3 patients with TKI resistance. Conclusion:Despite the limit of ourstudy, ourfinding highlights the high frequency of JAK2V617F/BCR-ABL double positivity as an important marker of resistance. So; the combination of JAK and TKI inhibitorsmightbe effective and potentiallybeguided by molecular monitoring of minimal residual disease1.

Keywords: chronic myeloid leukemia, tyrosine kinase inhibitors, resistance, JAK2V617F, BCR-ABL

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Authors: Murali Munisamy, Gauthaman Karunakaran, Mubarak Al-Gahtany, Vivekanandhan Subbiah, M. Manjari Tripati

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Background: Sodium valproate is a widely prescribed broad-spectrum anti-epileptic drug. It shows high inter-individual variability in pharmacokinetics and pharmacodynamics and has a narrow therapeutic range. We evaluated the effects of polymorphic uridine diphosphate glucuronosyltransferase (UGT1A9) metabolizing enzyme on the pharmacokinetics of sodium valproate in the patients with epilepsy who showed toxicity to therapy. Methods: Genotype analysis of the patients was made with polymerase chain–restriction fragment length polymorphism (RFLP) with sequencing. Plasma drug concentrations were measured with reversed phase high-performance liquid chromatography (HPLC) and concentration–time data were analyzed by using a non-compartmental approach. Results: The results of this study suggested a significant genotypic as well as allelic association with valproic acid toxicity for UGT1A9 polymorphic enzymes. The elimination half-life (t 1/2=40.2 h) of valproic acid was longer and the clearance rate (CL=937 ml/h) was lower in the poor metabolizers group of UGT1A9 polymorphism who showed toxicity than in the intermediate metabolizers group (t1/2=35.5 h, CL=1042 ml/h) or the extensive metabolizers group (t1/2=26. h, CL=1,302 ml/h). Conclusion: Our findings suggest that the UGT1A9 genetic polymorphism plays a significant role in the steady state concentration of sodium valproate, and it thereby has an impact on the toxicity of the sodium valproate used in the patients with epilepsy.

Keywords: UGT1A9, sodium valporate, pharmacogenetics, polymorphism

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Authors: Silpita Paul, Susanta Sadhukhan, Biswanath Maity, Madhusudan Das

Abstract:

Neural tube defects (NTDs) are one of the most common forms of birth defect and affect ~300,000 new born worldwide each year. The prevalence is higher in Northern India (11 per 1000 birth) compare to southern India (5 per 1000 birth). NTDs are one of the common birth defects related with low blood folate and Hcy concentration. Though the mechanism is still unknown, but it is now established that, NTDs in human are polygenic in nature and follow the heterogeneous trait. In spite of its heterogeneity, polymorphism in few genes affects significantly the trait of NTDs. Polymorphisms in the genes FOLH1 and MTHFR plays important role in NTDs. In this study, the polymorphisms of these genes were screened by bi-directional sequencing from 30 mothers with NTD babies as case. The result revealed that 26.67% patients had bi-allelic FOLH1 polymorphism. The polymorphism has been identified as p.Y60H and frequent to cause NTDs. The study of MTHFR gene showed 2 different SNPs rs1801131 (at exon 4) and rs1801131 (at exon 7). The study showed 6.67% patients of both mono- and bi-allelic MTHFR-rs1801131 polymorphism and 6.67% patients of bi-allelic MTHFR-rs1801131 polymorphism. These polymorphisms has been responsible for p.A222V and p.E429A change respectively and frequently involved in NTD formation. Those polymorphisms affect mainly the absorption of dietary folate from intestine and the formation of 5-methylenetetrahydrofolate (5 MTHF) from 5,10-methylenetetrahydrofolate (5,10- MTHF), which is the functional folate form in our system. Though the study is not complete yet, but these polymorphisms play crucial roles in the formation of NTDs in other world population. Based on the result till date, it can be concluded that they also play significant role in our population too as in control samples we have not found any changes.

Keywords: neural tube defects, polymorphism, FOLH1, MTHFR

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Authors: R. Komsa-Penkova, G. Golemanov, G. Georgieva, K. Popovski, N. Slavov, P. Ivanov, K. Kovacheva, S. Rathee, E. Konova, A. Blajev

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Leptin and PAI-1 are important cytokines and may play a role in the regulation of PCOS development. PCOS is frequently associated with obesity, high BMI index and consequently with increased risk of metabolic disorders. The aim of the present study was to evaluate PAI-1 levels, genetic influence of the carriage of 675 4G/5G polymorphism in PAI-1 gene and leptin as a marker of obesity in the development of PCOS. Methods: Genotyping in 84 patients with PCOS and PCO and 100 healthy control subjects to detect single nucleotide deletion 675 G in the promoter of PAI-1 gene. The present study provides evidence that SNP 4G in the PAI-1 gene is associated with early pregnancy losses in patients with polycystosis. Further to this, there is a correlation between leptin levels, PAI-1 levels and BMI in the patients with PCOS, which confirms the role of obesity as a risk factor for PCOS.

Keywords: carriage of 675 4G/5G polymorphism, PCOS, early pregnancy losses, PAI-1 gene

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Authors: I. Boroňová, J.Bernasovská, J. Kľoc, Z. Tomková, E. Petrejčíková, D. Gabriková, S. Mačeková

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Osteoporosis is a complex health disease characterized by low bone mineral density, which is determined by an interaction of genetics with metabolic and environmental factors. Current research in genetics of osteoporosis is focused on identification of responsible genes and polymorphisms. TNFRSF11B gene plays a key role in bone remodeling. The aim of this study was to investigate the genotype and allele distribution of A163G (rs3102735) osteoprotegerin gene promoter and G1181C (rs2073618) osteoprotegerin first exon polymorphisms in the group of 180 unrelated postmenopausal women with diagnosed osteoporosis and 180 normal controls. Genomic DNA was isolated from peripheral blood leukocytes using standard methodology. Genotyping for presence of different polymorphisms was performed using the Custom Taqman®SNP Genotyping assays. Hardy-Weinberg equilibrium was tested for each SNP in the groups of participants using the chi-square (χ2) test. The distribution of investigated genotypes in the group of patients with osteoporosis were as follows: AA (66.7%), AG (32.2%), GG (1.1%) for A163G polymorphism; GG (19.4%), CG (44.4%), CC (36.1%) for G1181C polymorphism. The distribution of genotypes in normal controls were follows: AA (71.1%), AG (26.1%), GG (2.8%) for A163G polymorphism; GG (22.2%), CG (48.9%), CC (28.9%) for G1181C polymorphism. In A163G polymorphism the variant G allele was more common among patients with osteoporosis: 17.2% versus 15.8% in normal controls. Also, in G1181C polymorphism the phenomenon of more frequent occurrence of C allele in the group of patients with osteoporosis was observed (58.3% versus 53.3%). Genotype and allele distributions showed no significant differences (A163G: χ2=0.270, p=0.605; χ2=0.250, p=0.616; G1181C: χ2= 1.730, p=0.188; χ2=1.820, p=0.177). Our results represents an initial study, further studies of more numerous file and associations studies will be carried out. Knowing the distribution of genotypes is important for assessing the impact of these polymorphisms on various parameters associated with osteoporosis. Screening for identification of “at-risk” women likely to develop osteoporosis and initiating subsequent early intervention appears to be most effective strategy to substantially reduce the risks of osteoporosis.

Keywords: osteoporosis, real-time PCR method, SNP polymorphisms

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