Bovine ephemeral fever (BEF) is an acute epidemic disease in cattle and water buffalo, spanning tropical and subtropical zones of Asia, Australia, and Africa. In recent years, BEF has been distributed in many provinces of Iran and caused economic losses. In this study the sequence encoding the G1 epitope of bovine ephemeral fever virus (BEFV) G glycoprotein was amplified by PCR, ligated into the prokaryotic expression vector, pMAL-c2X, and subsequently cloned into Rosetta strain of Escherichia coli. After being induced with Isopropyl β-D-1-thiogalactopyranoside, SDS-PAGE analysis showed that a protein of 58 kDa molecular weight, consistent with the expected molecular weight of maltose binding protein-G1 fusion protein, was expressed. Also, dot blot analysis confirmed that the expressed protein has specifically reacted with an anti-BEFV mouse serum. In conclusion, the results indicate that the recombinant G1 epitope expressed in this study could be used as a coating antigen to develop an ELISA for bovine ephemeral fever diagnosis.
Optimization of the condition for PCR-directed sequencing of microsatellites poly Adenine (A) length polymorphisms is more difficult and sensitive compared with other common sequences. Replication slippage may occur for polymerase enzyme during microsatellite amplification and direct sequencing of these PCR products will be challenging for heterozygote samples. So, the aim of this study is to introduce optimal condition for amplification of microsatellites poly (A) length polymorphisms such as variation of poly(A) at the 3′-end of the vitamin D receptor (VDR) gene. In this study firstly, we analyzed various types of microsatellite sequences by in silico method to select a short microsatellite sequence which has various alleles with a little length difference. Finally, VDR poly(A) microsatellite was selected. Amplification of this locus was optimized by applying hot-start PCR using gradients of temperature, DMSO, MgCl2, Taq polymerase, and primers concentration. Crush and soak method was used for DNA extraction of alleles in heterozygote samples from polyacrylamide gel. PCR products were analyzed by using single-strand conformation polymorphism and PCR-direct sequencing. Our observations showed that hot-start PCR was optimized in the concentration of 0.2 mM of primers, 0.8mM of MgCl2 and 1.75 unit of Taq polymerase in 25 μl PCR mixtures. These optimal conditions lead to remove Taq polymerase slippage and non-specific PCR product. Considering the possible errors in amplification and analysis of microsatellite, the presented method in this study could be an efficient strategy in analysis of these sequences.
The Autism Spectrum Disorders (ASDs) are common neurodevelopmental disorders estimated to affect 1 in 88 children. ASD is a complex condition, result of genetic, epigenetic and environmental factors. However, Genetic comopnent seems to play an important role. The loss/Gain of 1KB and more nucleotides, Copy number variation (CNV), is the significant genetic factor in the etiology of ASD. The CNV rate is reported as 10-15 in children with ASD. In this study we investigated CNV in 50 Iranian patients with sporadic Autism using High resolution Cytogenetics banding, MLPA, and Array-CGH techniques. All patients had additional features such as Intellectual Disability, Seizure, and Craniofacial anomalies. Two out of 50 (4) patients showed chromosome abnormality including 16p duplication (16p13.11-p13.3) and 15q deletion (15q11.2q13.1). MLPA using both sub-telomeic (p036 and p070) and Autism (p343 and p396) kits was done for 50 patients and CNV was detected in 5 (10) patients both in subtelomeric and interstitial regions, one overlapping the cytogenetic finding. Array CGH was performed for 15 patients most of whom had normal results with the Cytogenetic and MLPA techniques. Six out of 15 (40) patients using array-CGH showed significant CNVs including pathogenic (such as 15q24 Microdeletion), likely pathogenic (such as Xq28 Microdeletion) and Uncertain/ Could be Significant Ones (such as Xp22.33 Microduplication). We performed genotype-phenotype analysis and compared our results with other similar studies. To our knowledge, this is the first study on CNV in Iranian patients with ASD. We strongly recommend the investigation of CNV in patients with Autism with or without additional features. © 2016, Iranian Neurogenetics Society. All rights reserved.
Stroke is the third cause of death in the world after the cancer and cardiovascular disease. . It has been shown that stroke in Northeast of Iran occurs approximately one decade earlier than in Western countries. Reasons for the high stroke incidence among Persian are not yet clear. It is the first gene-association study in Persian population, which determines the relationship between 25 gene variations and stroke. This genes are involved in 3 cellular pathways and in Single nucleotide polymorphisms (SNPs) in miRNA genes. So we made a Genetic stroke databases with more than 1000 Stroke patients for this region. 25 gene polymorphisms were analyzed and registries in this database. Furthermore this database is part of INTERNATIONAL STROKE GENETICS CONSORTIUM . In the present study aimed to determine if there is any association between 22 genetic variations in protein-coding genes and also 11 polymorphisms in miRNA-coding regions. In this case-control study patients with ischemic stroke and subjects without any history of vascular diseases were recruited. All cases were diagnosed via computed tomography (CT) scans and magnetic resonance imaging (MRI). Genotyping was gen in cases and controls using Real-Time, PCR-RFLP and multiplex ARMS-PCR method. Statistical analyses were performed using SPSS v.21 software and a pvalue< 0.05 was considered significant.
The objective of the present study was to evaluate the genetic structure and diversity of 16 generations of native fowls of West Azerbaijan Province, Iran using the pedigree data collected between 1994 and 2012. Inbreeding coefficient (F), pedigree completeness index (PCI), parameters of the probability of gene origin and other parameters related to the genetic structure were estimated using the pedigree data of 51125 chickens. The mean PCI for up to 3 and 16 generations was 0.798 and 0.365 in the whole population, and 0.951 and 0.448 for the inbred population respectively, showing the sufficient depth of the pedigree and higher amounts of PCI for the inbred population. The total number of animals in 16 generations was 51125 chickens (F=3.07%); and the number of sires and dams was 1352 (F=2.247%) and 6863 (F=2.71%), respectively. The number of inbred chickens, sires, and dams in the population was 40751, 634, and 4429, respectively. Despite the large number of inbred animals in the population, the inbreeding average was not so high in different generations. Thus, it may be concluded that there is a suitable potential for line-crossing to help with maintaining genetic diversity in the population. The highest percentage of line-crossing was observed in the 6th (15.2%), 8th (13.81%) and 10th (10.71%) generations and its lowest percentage was in the 12th generation (1.59%). All parameters of the probability of gene origin had a decreasing trend in the studied generations, and the value of the non-founder genome equivalents was higher than the effective number of founders after base generations (after the third generation). Therefore, genetic bottleneck and random genetic drift may be the main causes of genetic diversity loss in this population.
One of the important mechanisms utilized by the plant growth promoting Rhizobacteria (PGPR) is lowering of ethylene levels by the 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase enzyme. The mechanism facilitates plant growth and development in the stress conditions. PGPR containing ACC deaminase can promote plant growth under stress and overcome the harmful effects. The study aimed to screen and identify the ACC deaminase producing rhizobacteria isolated from the soils of different regions in Iran. Fifty-seven isolated strains of rhizobacteria were cultured in the M9 medium with two different nitrogen sources (contains ACC and NH4Cl), in two replications. The control treatment was nitrogen free M9 medium. In this study, canola seeds, Sari Gol cultivar, were also inoculated with the ACC deaminase producing rhizobacteria for evaluating their effects on the shoot and root elongation during 5 days germination based on a completely randomized design with three replications. Finally, the best ACC deaminase producing rhizobacteria was identified by the sequencing of 16s rDNA gene. Results showed that just 8 isolated rhizobacteria strains (out of 57) produced ACC deaminase, of which Cma3 and Wah3 had the greatest effects on the stem and root length, respectively. Cma3 and Wah3 were identified as Klebsiella and pseudomonas genus, respectively. The identified strains can be useful for the crop growth in stress conditions.
ipaD gene is amongst key genes in Shigella invasion, known as an antigen. PA20 protein of Bacillus anthracic can be separated from native PA protein during the infection and can be found in the serum of the infected patients and make it possible to diagnose the disease. CTxB, a known immune adjuvant, enhances the immunogenicity and is mainly used for the production of recombinant vaccines as a booster for immunization with antigen. In order to produce a vaccine with the immunogenic properties for the diseases, shigellosis and anthrax, different combinations of three ipaD, PA20 and CTxB genes were studied by bioinformatic tools in terms of their stability and similarity to the native proteins that are essential for their accurate performance. The aim of the present study was to introduce the best combination of the three antigens (ipaD, PA20 and CTxB) such that, when they are fused together, have a minimum effect on their structure and activity. Secondary and tertiary structures of proteins derived from different combinations of two or three proteins of ipaD, PA20 and CTxB were studied by MODELLER and PSIPRED software and the stability and lifespan of the fused genes were surveyed using ProtParam software. Analysis of data obtained from different combinations of ipaD, PA20 and CTxB genes by PSIPRED, ProtParam and MODELLER software showed that the best triple combination was PA20-CTxB-ipaD and the best of dual combinations was PA20-ipaD. When a protein combines with other proteins, the three-dimensional structure and physicochemical characteristics may dramatically change. According to the results of the present study, PA20 showed a strong stability, while ipaD showed a poor stability. Furthermore, all proteins in PA20- CTxB-ipaD combination had maintained their stability and this fusion can be used furthermore for vaccine design.
Hearing loss (HL) is the most frequent sensory defect present in 1 of every 500 newborns. In developed countries, at least 50% of cases are caused genetic factors, most often resulting in nonsyndromic HL (70%), which is usually autosomal recessive (80%). To date, fifty genes associated with autosomal recessive non-syndromic hearing loss (ARNSHL) have been reported. The aim of this study was to determine the prevalence of mutations in four genes (GPSM2, MSRB3, SLC26A5, and GRXCR1) in Iranian deaf population with ARNSHL. These genes were mostly reported in our neighboring countries. One hundred unrelated Iranian families segregating ARNSHL with at least two affected siblings were subjected to our study. We used homozygosity mapping to identify regions of autozygosity-by-descent using three flanking or intragenic short-tandem repeat (STR) markers for mentioned genes. If a family showed linked pattern to our selected STR markers, then direct sequencing was performed. Seven families were linked to GPSM2, seventeen families were linked to GRXCR1, five families showed linkage to SLC26A5, and sixteen families linked to MSRB3. To confirm the mutation in linked families, direct sequencing was performed, however, after analyzing the sequencing results, no mutation could be detected in either of the families. Our data showed that mutation in the studied genes, GPSM2, MSRB3, SLC26A5, and GRXCR1, is not prev
microRNAs are highly conserved small non-coding regulatory RNAs that involve in post transcriptional regulating of gene expression during different cellular mechanisms. Aberration of miR-218 expression during tumorigenesis of different solid tumors has been reported by numerous studies. In current systematic review article, by using the terms “miR-218” and “cancer” we first searched for English language articles in the PubMed database, published from 1993 to April 2014. Then by a comprehensive review of related articles, we provided some new insights that highlight novel features and functions of miR-218 in initiation and progression of solid tumors. The majority of these studies propose a tumor suppressing role for miR-218 considering the fact that it is significantly down-regulated in tumor tissues compared with normal specimens. Despite accumulating body of evidence regarding tumor suppressor functions of miR-218 in solid tumors; more intensive reviewing about available miR-218 recent original studies and interpretation of existing data, revealed the multifunctional role of miR-218 in these kinds of malignancies by targeting different corresponding target genes. Take all together, MiR-218 targets different cellular processes in cancer cells and its expression pattern is in an important association with various states and features of tumors. It seems that miR-218 can increase the speed of cell cycle and cell division in lower sample grades and along with progression of cancer cells it's function changes to stabilization the cancer cells and not allowing them to invade. thats why it often shows up-regulation in lower grades and down-regulation in metastatic phase. Therefore, it seems of great importance to check samples stage, grade, lymph node metastasis status and other tumor features before evaluation of miR-218 as a prognostic or diagnostic biomarker.
Normal sex differentiation is the result of long ways and different steps that consecutively take place under the control of relevant enzymes resulting in a normal female or male. Any effective agent and excess or deficit of the relevant enzymes result in the disorder of sex development. The main factors that affect sex development include chromosomal aberration, gonadal dysgenesis, and hormonal effects. Normal sex differentiation and development have been described in the first chapter in the Genetics in the Third Millennium (Vol. 13, No. 1). The disorders of sex development due to chromosomal aberration and gonadal dysgenesis have been delineated in the second (Vol. 13, No. 2) and third chapters (Vol 13, NO. 3). In the fourth chapter, we present and discuss the disorder of sex developments due to hormonal effects.
Genetic distance is a traditional measure of divergence among populations (or species) which can be used to infer whether populations are panmictic, or sorting as reproductively independent (more or less) units. There are many different quantitative approaches to measure genetic distances. The concept of genetic distance was developed as early as 1953 by Sanghvi in a study of the genetic differentiation of human populations and was later refined by others (4), however, a distance measure that is appropriate for studying protein evolution was developed only after electrophoretic studies became popular. In recent years, statistical properties of various distance measures have been studied. Since Cavalli-Sforza and Edwards (3) published their work, many geneticists were interested in this problem and various measures of genetic distance were proposed. Cavalli-Sforza & Edwards (4) attempted to relate their distance measure to the evolutionary changes of gene frequencies. Latter (14) proposed several measures that are closely related to Wright's (33) fixation index. Nei (17,18) proposed a genetic distance measure with which one can estimate the number of gene or codon substitutions Abstrcat: The aim of the present study was to find the best or optimum topology of six chicken populations, which were genotyped based on nine highly polymorphic microsatellite markers. To reach this goal, different genetic distances based on infinite allele model (IAM), stepwise mutation model (SMM) and drawing phylogentic trees on un-weighted pair-group method using arithmetic averages (UPGMA) method were compared. For conducting this research, 16 DNA samples from each Naked Neck (NN), White Silkies (WS), Commercial Layer (CL), Commercial Broiler (CB), Giriraja (GR) and Desi (DS) populations were genotyped for this study. Some of the IAM and SMM based genetic distances showed the same topology under UPGMA. Estimates of different genetic distances revealed relatively close genetic similarity between NN and GR. In the UPGMA-based phylogenetic trees constructed from different genetic distances, four major clusters obtained first, NN and GR; second, NN, GR and CB; third, DS and CL; and forth, WS in a single cluster. The percentage occurrence of each group among genetic distances was 100, 100, 66.7 and 55.6, respectively. The correlation coefficients obtained by Mantel’s test ranged from 0.470 to 0.997 between different genetic distances. In conclusion, by applying this procedure, it is possible to get proper topology using different genetic distances. Keyword: Phylogenetic Tree; UPGMA; Chicken; Microsatellite Repeats Applying Different Genetic Distances and UPGMA Method to Find the Best Topology in Chicken Populations Using Microsatellite Markers
Abstrcat: Acute myeloblastic leukemia is the most frequent cancer in adults and the patients have a wide range of sub-classes. HLA DR negative cases represent one of the major immune-phenotypic classes. HLA-DR is routinely used to distinguish acute promyelocytic leukemia (APL) from other AML subclasses based on the immunophenotype. The TGF-b signaling pathway, as a critical regulator, guides different vital and sometimes opposite aspects of cell processes, including cell proliferation, apoptosis, differentiation quiescence, and malignancy expansion. A review of previous studies regarding the role of TGF-b in human malignancies made us interested in the evaluation of the role of the TGF signaling pathway in leukemia. We evaluated TGF-b / TGF-bRII fluctuant in “HLA-DR negative AML”, at expression level in bone marrow and peripheral blood samples. Forty-six patients were consecutively diagnosed with HLA-DR negative AML by flow cytometry, morphology, cytochemistry, and molecular analysis. By using Ficoll density centrifugation, mononuclear cells were isolated from the peripheral blood and bone marrow from both patients and controls; then, TGF-b, TGFbRII, and ABL1 genes were amplified by quantitative Real Time PCR. The results revealed that the TGF-b expression level was not different between patients and controls while TGF-b R was higher in patients than control cases; the expression of TGF-b and TGF-bRII was significantly lower in non M3-AML (M0, M1, M2) than M3-AML (APL) (P<0.05). We conclude that despite the conventional role of the TGF-b/TGF-bRII pathway to induce quiescence, anther role was defined for leukemic cells expansion in HLA-DR negative AML. Currently we are studying this pathway to find the downstream tar get of leukemogenesis activity. Keyword: HLA-DR negative; Leukemia, Myeloid, Acute; Transforming Growth Factor beta; TGF-bRII
