Journal of Cell and Molecular Research

Online ISSN: 2008-9147
Publications
Total phenolic and flavonoid contents of T. transcaspicus in different stages of sampling.
Article
Antioxidant activities protect the cell against oxidative agents that are constant metabolic by-products. The aim of this study was to investigate the relationship between harvesting time of Thymus transcaspicus and its antioxidant activities. The plant samples were harvested 5 times in different growth phases from 17 April to 22 July 2008, and its antioxidant activity was studied using the ferric reducing antioxidant power (FRAP), 1,1- diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging activity, and β-carotene bleaching (BCB) assays. The results of FRAP assay indicated that the reduction activity of the plant was in the highest level in stage 5 of sampling. The result of DPPH assay showed that the crude extract of the plant was more capable of DPPH radical scavenging in stage 2. The highest level of gallic acid and quercetin in the crude extract of T. transcaspicus was determined as 85.29 ± 6.22 mg and 18.88 ± 0.9 mg in stage 2, respectively. Therefore, stage 2 was the optimum time to harvest the T. transcaspicus.
 
Article
Trehalose is the alpha, alpha-1,1-linked glucose disaccharide. Its metabolism is found in a wide variety of organisms and is seen as evolutionary old. Trehalose metabolites are however present at only very low concentrations and their role in plants are not understood. The physiological effects of 100 mM trehalose on growth and carbon allocation in seedlings are characterized in this paper. Trehalose feeding to Arabidopsis thaliana elicits strong responses. On 100 mM trehalose, seedlings germinate and extend cotyledons but fail to develop primary leaves. The primary roots do not grow beyond 2-3 mm and there is not any starch in root tips. In light, growth arrest on 100 mM trehalose can be rescued by exogenous supply of metabolisable sugar. Trehalose feeding results in anthocyanin accumulation and chlorophyll reduction. Trehalose causes cells of the root extension zone to swell and lysis. Trehalase expression analysis showed, WT seedlings grown on trehalose have 10-fold induced AtTRE1 expression compared to sorbitol treatment.
 
RAPD profiles of genomic DNA from P. Putida . Control (top) and F. assafoetida latex treatment (down). M: 100 bp DNA ladder (100-3000) and 1 to 18 are primers as showed in table 1. 
RAPD profiles of genomic DNA from P. Aeruginosa . Control (top) and F. assafoetida latex treatment (down). M: 100 bp DNA ladder (100-3000) and 1 to 18 are primers as showed in table 1. 
RAPD profiles of genomic DNA from X. compestris . Control (top) and F. assafoetida latex treatment (down). M: 100 bp DNA ladder (100-3000) and 1 to 18 are primers as showed in table 1. 
RAPD profiles of genomic DNA from S. aureus . Control (top) and F. assafoetida latex treatment (down). M: 100 bp DNA ladder (100-3000) and 1 to 18 are primers as showed in table 1. 
RAPD profiles of genomic DNA from B. subtilis . Control (top) and F. assafoetida latex treatment (down). M: 100 bp DNA ladder (100-3000) and 1 to 18 are primers as showed in table 1. 
Article
Asafoetida is the dried latex exuded from the living underground rhizome or tap root of Ferula assafoetida. Antibacterial characteristic of asafoetida was shown using the circular zone diameter of bacterial growth inhibition by disk-diffusion method on two gram positive and three gram negative bacteria. Then, the bacterial genomic DNA damage, induced by F. assafoetida latex, was demonstrated using the comparison of random amplification of polymorphic DNA profiles generated by polymerase chain reaction of control and treated bacterial genomes. The results showed that the number of primers that produced bands in each bacterium were higher in control samples compared to those treated with asafoetida. This and the absence or presence of bands between controls and treatments confirm rearrangements and DNA damage in the priming binding sites of bacterial genome.
 
Proposed patterns for genotyping of the observed bands.
SSCP analysis of the 275 bp fragment of Leptin gene (exon 3). Lanes from left to right (1-18): ABDE, ABCF, ABF, ABC, ABCF, ABE, CC, AC, AB, AB, ABCF, ABC, ABE, AB, ABE, AC, ABCF, Ladder (M50).
, were observed and only two of them were homozygous. Distributions of the alleles of Leptin gene in different animal groups and other diversity measurements are presented in table 2. The following equation was used to calculate genotype frequency of the A/B genotype:
Article
Identification of genes affecting energy balance, milk yield and feed intake is an interesting area of researches in animal breeding. Leptin gene polymorphism is associated with key economic affair. Considering rich resources for animals, in our country, accomplishing a few assays to identify a gene that controls her traits with molecular genetics, and identifying the candidate genes in sheep breeds using DNA test can greatly help to her breeding progress. For analyzing Leptin gene polymorphism and its association with growth traits in Kermani sheep, blood samples of 120 sheep of both genes rearing at breeding centre of Shahre Babak were taken. In addition growth traits were measured. PCR was performed to amplify 275 bp fragments of exon 3 from Leptin gene. Then Single Strand Conformation Polymorphism (SSCP) of PCR product was performed and Leptin band patterns (genotypes) were obtained using acrylamid gel and silver staining. For Leptin gene 10 genotypes including A/A, C/C, A/B, A/C, A/B/C, A/B/E, A/B/F, A/C/F, A/B/D/E and A/B/C/F were obtained. The results of this study showed that the growth traits are significantly affected by the genotypes. Accordingly, A/B/E, A/C, A/B/C/F and A/B/C/F genotypes had higher body weight at 3, 6, 9 and 12 months of ages respectively. The animals with A/B, A/B, A/B/F and A/B/D/E genotypes had the smallest body weight at 3, 6, 9 and 12 months of ages respectively. It is suggested that polymorphism in Leptin gene loci can be used as a selective criterion to improve growth traits in Kermani sheep.
 
The O-shaped rings as punched from rabbit pinna in culture medium.  
Morphology of SLCs in fifth passage (scale bar: 50 μm).  
Growth rate analysis of SLCs grown in media containing various percentages of FBS, as examined by counting the cells at different time points (data are shown as Mean+/-SD).  
Karyotype analysis of SLCs, derived from male New Zealand white rabbit pinna at passages 65.
Article
Regeneration is a biological phenomenon, which takes place via two main mechanisms: first, dedifferentiation of mature cells followed by their differentiation into functional new cells and second, activation of endogenous somatic stem cells for regeneration of damaged or lost tissues. One of the best examples of healing process in mammals is the regeneration of damaged pinna in rabbits by blastema tissue. The aim of present study was to investigate culture requirements, proliferative properties and expression of some stemness factors in cells derived from regenerating blastema tissue obtained from rabbit pinna in vitro. The regenerating tissues were obtained from male New Zealand white rabbits by double punching of the pinna and cell culture conditions were set to derive and enrich the self renewing cells for further characterisation. The cells were subjected to survival and growth examinations in vitro, and expression of several stemness factors was studied in these cells using reverse transcription polymerase chain reaction (RT-PCR). Results revealed that the derived cells are rather immortal, as they have been growing for more than 120 passages in culture up until this report. Furthermore, RT-PCR and flow cytometry analyses showed that these cells express a number of stemness related genes including Oct4 and Sox2. In conclusion, in this study, stem like cells were derived from blastema tissue of rabbit ears for the first time, showing great self renewing capacity, which provides a suitable in vitro model for regeneration studies. Moreover, they could be considered as a good source of stem like cells for future applications.
 
Article
Microglia, the sentries of the brain, is highly implicated in neurodegeneration as in neuroprotection. Chronic microglial activation endangers neuronal survival through the release of various potentially neurotoxic mediators including Nitric Oxide (NO). Thus, negative regulators of microglial activation have been considered as potential therapeutic candidates to target neurodegeneration, such as those in Alzheimer's and Parkinson's diseases and even in chronic epileptic syndromes. Bromelain, a mixture of cysteine proteases, derived from pineapple stem, has been reported to have anti-inflammatory and immunomodulatory effects. Neonatal rat primary microglial cells were isolated from the brain according to the Floden's method. The purity of the cultures was determined by immunostaining with an OX-42 antibody which showed a purity greater than 95%.The activation profile of microglia was investigated by determining the effects of Bromelain (1, 10, 20, 30, 40 and 50 µg/ml) on the level of neurotoxin, NO, mitochondrial activity and morphological changes in treated microglia with lipopolysaccharide (LPS) (1µg/ml), as an endotoxin. Our results showed that pretreatment of primary rat microglia with bomelain (30 µg/ml), decreased the production of NO induced by LPS (1µg/ml) treatment in a dose-dependent manner, which prevented the deramification of microglia and its phagocytic morphology. Moreover, bromelain does not show cytotoxicity at any of the applied doses, suggesting that the anti-inflammatory effects of bromelain are not due to the cell death. In conclusion, Bromelain reduces the NO synthesis in vitro by potentially exerting its anti-inflammatory effects. Bromelain naturally found in pineapple stem, can be considered as a useful agent for neuroprotection and alleviation of symptoms in neurodegenerative diseases.
 
Effect of GA 3 on photosynthetic and non photosynthetic pigments of tobacco leaves (NT= non transgenic, T= transgenic). Similar letters represent no significant differences (P<0.05).
Article
Transgenic and non transgenic Nicotiana tabacum L (cultivar Wisconsin) containing Ri T-DNA were treated with 0, 0.2 and 0.4 mgL -1 GA 3 in Murashig and Skoog medium. Some physiological parameters including shoot length, leaf area, number of auxiliary bud, fresh and dry weight, number and length of trichomes were measured. Shoot length, fresh weight and dry weight were increased but number of trichome did not change by GA 3 treatment. Chlorophyll, carotenoid and anthocyanin pigments of leaf were decreased. Auxin and gibberellic acid content of leaf and root were also measured. Exogenous GA 3 increased root auxin in the transgenic plants while it did not change in shoot. GA 3 treatment increased gibberellin content in both of root and shoot.
 
RAPD profiles of safflower varieties and saffron samples amplified with RAP5 on 1.2% agarose gel (0: Size marker (100 bp), lanes 1-7: Safflower varieties (1: IL-111, 2: 2819, 3: 279, 4:K.W.3, 5: K.W.6, 6: K.W.16, 7: 295), Lanes 8-12: Saffron samples; (8: Ghaen, 9: Gonabad, 10: Barakuh Gonabad, 11: Torbat Heidariieh, 12: Science and Technology Park)  
PCR amplification of safflower varieties using SAF-L40 on 1.2% agarose gel (M: Size marker (100 bp), lanes 1-7: safflower varieties (1: IL-111, 2: 2819, 3: 279, 4:K.W.3, 5: K.W.6, 6: K.W.16, 7: 295). Lanes 8-12: Saffron samples (8: Ghaen, 9: Gonabad, 10: Barakuh Gonabad, 11: Torbat Heidariieh, 12: Science and Technology Park).  
PCR amplification of safflower and saffron using SAF-L70 on 1.2% agarose gel (lane1: size marker (100 bp), lane2-PCR of saffron DNA (all of saffron samples) by SAF-70 primers, lane3: PCR with DNA mixture of safflower varieties by SAF-70 primers.  
Article
Saffron (Crocus sativus) is the most valuable and indigenous crop in Iran. The stigmas of flower are used as a popular natural flavouring, colouring and medicinal agent. However, the market suffers from frauds in this plant such as mixing with safflower petals due to high profit. Identification of these frauds with conventional and biochemical methods is difficult and low sensitive. Therefore, application of molecular markers such as random amplified polymorphic DNA (RAPD)/sequence characterized amplified regions (SCAR) is being considered as an alternative. In this study, DNA was extracted from dry stigmas of 5 Saffron accessions and dry petals of 7 safflower cultivars. RAPD reactions with ten 15-mer random primers resulted in two specific monomorphic bands (500 and 700 bp) for safflower, while they were absent in saffron accessions. PCR analysis with specific SCAR primers amplified two specific bands (414 and 589 bp) for safflowers in different combinations of saffron stigmas and safflower petals. This was the case with very low rates or 1% of safflower. Therefore, this method seems to be suitable for fraud identification of safflower petals in commercial saffron samples.
 
The amount (ppm) of different silican compound in the extract of Equisetum telmateia stem and leave, measured by the Atomatic Absorption Spectrometry technique. 
Article
Equisetum telmateia (Equisetaceae) seems to have anti inflammatory and antioxidant properties. In the present study, the neuroprotective effects of organic and inorganic silica were investigated on spinal cord alpha motoneuron of rats after injury of sciatic nerve. After highly compression of sciatic nerve in 42 Wistar rats, the injured rats were divided into sham (n= 6) and two experimental groups which each were divided into 3 subgroups (n= 6). The first subgroups received 3, 6 or 9 injections (15 mg/kg/injection, ip) of horse tail extract and the second subgroups received 3, 6 or 9 injections (6 mg/kg/injection, ip) of sodium meta silicate, respectively. The first injection was made after sciatic nerve injury and the others by 72 hours intervals. After a month, the rats were sacrificed and their spinal cord lumber segment sampled, processed for histological preparation and analyzed stereologicaly (the dissector technique) for estimation of numerical density of alpha motoneurons. The results showed significant decrease in the numerical density of alpha motoneurons in shams (p< 0.05) and no significant differences between experimental and control groups. This may suggest the neuroprotective effects of silica on the survival of alpha motoneurons.
 
Bone marrow smear from control rat. PCE are stained light purple.
Bone marrow smear of treated rat. MNPCE is at the center.
Article
Due to the wide range of applications for ozone and its increasing use for medical and industrial purposes, studying its effects has become a very important line of research. The ozone has been suspected to be a carsinogen. Because of the increasing use of ozone, the human could be more and more exposed to this gas. In this study the effects of ozone inhalation on chromosomes and its clastogenic consequences have been investigated using in vivo micronucleus assay in bone marrow cells of treated rats. Animals were treated for 6 hours a day at 3 ppm of ozone during 10 consecutive days. The micronucleus assay was performed immediately and 11 days after the last exposure. The frequency of micronucleated polychromatic erythrocyte of bone marrow (MNPCE) increased in both groups compared to the control. Such increase confirmed the clastogenic effects of ozone. The elevated frequency of MNPCE did not decrease after 11 days of the last ozone exposure. Results indicate that ozone inhalation could induce persistent chromosomal damages even to bone marrow cells which were not in direct contact to it. Also, once more, the results confirmed the usefulness of the micronucleus assay in toxicological studies.
 
Article
Study of the different aspects of protection against the exposure of ionizing radiation has always been an active area of research. High cost and toxicity of radioprotective drugs have limited their use. So, search for new drugs with a high degree of protection and lower cost and side effects seem a necessity. In this study radioprotective effect of aqueous as well as alcoholic extracts of the Mann of Cotoneaster nummularia( Shirkhesht), regarding their high accessibility and possibly low side effects, against 2 Gy Gamma irradiation, was analyzed using micronucleus assay on bone marrow cells of male mice (Balb/c). Different doses of 250, 500, 1000 mg/kg/BW for aqueous and 3750, 7500, 15000 mg/kg/BW for alcoholic extract of Shirkhesht were administered IP for five constitutive days prior to 2 Gy gamma irradiation. The result compared with the known radioprotective effect of vitamin E after the same treatment schedule. High frequency of micronucleus was observed in non treated gamma-exposed mice, which represented the clastogenic effect of irradiation. Vitamin E, aqueous and alcoholic extracts of Shirkhesht treated mice represented a 5.56, 3.32 and 2.1 times decrease in the gamma-induced micronucleus frequency respectively. The data suggest a radioprotective effect of shirhkesht compared to vitamin E.
 
Article
Phosphoenolpyruvate Carboxykinase, encoded by the pepck gene, plays an important role in gluconeogenesis. It also seems to be important in metabolism of nitrogenous compounds in developing seeds of legumes, including amides and ureides which are then transformed into amino acids, necessary for the synthesis of storage proteins. In this research, pepck gene expression in mRNA level, in different genotypes of chickpea (Cicer arietinum L.), was determined. Two low protein genotypes (MCC291 and MCC373) and two high protein genotypes (MCC458 and MCC053) out of 20 chickpea genotypes were selected. Total RNA were extracted through different stages of seed development, and the expression of the pepck gene was estimated by semi-quantitative RT-PCR. The results of the RT-PCR showed that two isoforms of this gene are expressed in high protein genotypes, whereas in the low protein genotypes, the expression of these isoforms was not obvious. Also this method showed a differential expression of pepck gene in different stages of flowering and seed development. pepck gene is expressed in higher levels during the sheet formation and developing seeds compared to the flowering and seed formation stages. Probably, the differential expression of pepck gene is related to its possible role in metabolism of seed components, particularly in determination of the protein content of chickpea seeds.
 
a. Transverse section through kidney paranchyme on day 13 of gestation that collagen indicated no reaction in glomerular primordium (arrow heads) as well as rudimentary tubules (arrows). b. Other transverse section during glomerular development on day 14 of gestation. Although glomerular structure have been completed, no reaction was observed in any area except for parenchyma vessles. 
a. Transverse section of glomerulus on day 15 of gestation, The first reaction was observed in basement membrane of glomerulus of cortical regions (arrow heads). b. Sections of glomerulus on 18 day of gestation. In this image it have been showed labeling in both basement membrane of cortical glomerulus (arrow heads). 
a. Transverse sections through glomerulus on 5 th postnatal day, arrowheads refer to strong reaction in glomerular basement membrane. b. Glomerulus on 10 th 
Article
Basement membrane of glomerular mesangium (BMG) is a thin membrane which helps to support the capillary loops in a renal glomerulus and type IV collagen is require for complete BM formation during glomerulogenesis. In this investigation specific antibody of type IV collagen has been used in light microscopy to study development of BMG of the embryonic and postnatal mouse glomerular mesangium. In this study, 20 female Balb/C mice were selected randomly and finding vaginal plug was assumed as day zero of pregnancy. 12 pregnant mice were sacrified by cervical dislocation in one of gestational days 13-18, their fetuses were fixed and serially sectioned. Immunohistochemical Study for tracing of collagen type IV in BMG was carried out. The same processes were carried out for kidneys preparation of pups on 5, 10, 15 and 20 days after birth (2 mothers for each day). The result of the present study revealed that collagen IV reaction was weak on day 15 of gestation. The amount of collagen increased continuously until next days of fetal life and primary of 10 days postnatal in BMG. After this period, collagen IV showed no significant change in newborns. These data indicate that collagen IV appears just during the glomerular vasculogenesis and because of continuity and glomerular endothelial cell differentiation, type IV collagen, is the major structural protein in BMG, have been implicated in these processes. Keywords: collagen IV, glomerular basement membrane, kidney, mouse
 
RWC in control and drought-treated plants. PEG 100, 200 and 400 g/l are equal to-0.15,-0.49, and-1.76 MPa water potential, respectively. Data are means ±S.E. of three replicates. Treatments with the same lowercase letters were not significantly different based on mean comparison by Duncan's method at P < 0.05.
Article
Osmotic stress is one of the major factors that significantly reduce yields in dry areas. Plants respond to this abiotic stress at physiological and molecular levels. Many genes are induced under stress conditions by transcription factors. Dehydration responsive element binding (DREB) protein is a subfamily of AP2/ERF transcription factors which control expression of many osmotic stress-inducible genes. In this study, 21 days old seedlings of Sardari cultivar, dry farming bread wheat transferred into hydroponics culture using Hoagland solution. Osmotic stress treatments performed with adding 100, 200 and 400 g/l poly-ethylene glycol 6000 to hydroponics culture to obtain –0.15, –0.49, and –1.76 MPa water potential, respectively. After the seedlings were withered and colorless, relative water content, dry weight, and photosynthesis measured. In addition, RTPCR, and cDNA sequencing carried out. Molecular analysis of DREB translated protein sequence performed by DNAMAN, BLASTN, Pfam and PROSITE software. Results showed that osmotic stress decreased relative water content, root and shoot dry weight and net photosynthesis rate in comparison to control, significantly (P < 0.05). Sequence alignment indicated 98% homology with other Triticum aestivum DREB protein mRNA. There was an AP2 domain in the translated protein with three -sheets and one -helix and contains the Val14 and Glu19 amino acids. An EST Sequence deposited in NCBI GenBank database with the accession number of ES466900.
 
The list of primers sequence 
Article
Schizophrenia is a sophisticated mental disability which has affected nearly1.1% of people all over the world. According to recent researches, the key proteins triggered in the immune system are cytokines which might also be taking part in the pathogenesis of schizophrenia. The aim of this study was to evaluate the relationship between the-1082G/A and +874T/A polymorphisms of IL-10 and IFN-γ genes, respectively, in patients with schizophrenia. Total of 94 schizophrenic patients and 97 individuals as control samples were enrolled in this study. All samples were genotyped by amplification mutation refractory system-polymerase chain reaction (ARMS PCR) for candidate SNPs in IFN-γ and IL-10 genes. No significant association was found among various genotypes of IFN-γ and IL-10 in selected SNPs with risk of schizophrenia. As well as there was no significant variation in allelic frequency of IFN-γ and IL-10 genes with the risk of disease. These data suggest that the-1082G/A of IL-10 and +874T/A IFNγ genes are not involved in the development of schizophrenia risk. To validate of this data, suggesting more studies in diverse populations with larger sample size.
 
Article
Wound healing is a complex biological process in which many molecules, including microRNA molecules, play an essential role in its regulation. It is well-established that reducing miR-155 expression can accelerate wound healing. This study investigated the effect of using nanoparticles loaded with vitamin C on miR-133, collagen I, and III expressions. In this study, first, nanoparticles of albumin protein were produced and then loaded with vitamin C. 3T3 mouse fibroblast cells were affected by these nanoparticles, and cell behavior was investigated to evaluate the toxicity and appropriate doses. In addition, the expression of collagen I and III genes was studied. The results showed that nanoparticles containing vitamin C in 20 µg/ml concentration had a positive effect on collagen I and III expressions compared to the control group. Moreover, we observed a decrease in the expression of miR-133 in comparison to the control group. Therefore, according to the results of this study, it can be argued that nanoparticles containing vitamin C can significantly decrease the expression of the miR-133 gene and lead to collagen I and III gene overexpression in fibroblasts cells, which is directly effective in wound healing.
 
Article
Applying microorganism in oil recovery has attracted attentions recently. Surfactin produced by Bacillus subtilis is widely used industrially in a range of industrial applications in pharmecutical and environmental sectors. Little information about molecular mechanism of suffactin compound is available. In this study, we performed promoter and network analysis of surfactin production genes in Bacillus subtilis subsp. MJ01 (isolated from oil contaminated soil in South of Iran), spizizenii and 168. Our analysis revealed that comQ and comX are the genes with sequence alterations among these three strains of Bacillus subtilis and are involved in surfactin production. Promoter analysis indicated that lrp, argR, rpoD, purr and ihf are overrepresented and have the highest number of transcription factor binding sites (TFBs) on the key surfactin production genes in all 3 strains. Also the pattern of TFBs among these three strains was completely different. Interestingly, there is distinct difference between 168, spizizenii and MJ01 in their frequency of TFs that activate genes involve in surfactin production. Attribute weighting algorithms and decision tree analysis revealed ihf, rpoD and flHCD as the most important TF among surfactin production. Network analysis identified two significant network modules. The first one consists of key genes involved in surfactin production and the second module includes key TFs, involved in regulation of surfactin production. Our findings enhance understanding the molecular mechanism of surfactin production through systems biology analysis.
 
List of characters used for chromosome analysis
Dendrogram showing the phonetic relationship among the studied localities of Mus musculus, constructed using the matrix of symmetrical indexes similarities with UPGMA. 
Dendrogram showing the phonetic relationships among the populations of Mus musculus captured from different localities, constructed using the matrix of karyotype similarities with UPGMA All specimens from central and eastern Iranian house mouse are separated by the greater phenetic distance and they are placed in second cluster. 
Article
Cytotaxonomy is a branch of cytogenetics, devoted to the comparative study of karyological features for systematic and evolutionary purposes. Surely, awareness of chromosomal characters increases our knowledge in different fields of studies. In this study, cytogenetic analyses were performed in 92 Mus musculus specimens from 26 localities in Iran. Cytogenetic characteristics of the house mouse, Mus musculus, in Iran show that the chromosome number is 2n=40 and the arm number is NF=40. The karyotyping results indicated the presence of 20 Acrocentric (A) chromosome pairs. The L/S (r ratio) was between 2.0621 and 4.5862. The length of shortest chromosome, length of longest chromosome and mean of chromosomal length in different populations were between 2-3.58, 6.07-7.01 and 3.43-5.05 (μm), respectively. The results showed two distinct karyotypic formulae, namely cytotype B and cytotype C. Asymmetry indexes (AI, DI, As%, A, A2, A1 and Syi%) in all population except Birjand and Khash showed symmetry in chromosomes. In clustering methods using the matrix of symmetrical indexes similarities, four clusters were revealed, one for specimens of central and east of Iran, the second cluster for specimens from south and west of Iran, the third cluster was related to the eight specimens of Birjand and finally, the fourth cluster for two specimens of Khash locality.
 
Article
Genetic diversity is one of the three levels of biodiversity. The aim of present study was to compare levels of genetic polymorphism between wild Bream populations using seven microsatellite loci. Genetic diversity was investigated by studying samples collected from two regions, the coast of Chamkhale and Bandaranzali of Gilan province. A total of seven microsatellite loci (MFW7, MFW26, Mcs1EH, Rser10, Bl1-153, Bl2-114 and IC654) were used. The average number of alleles in Chamkhale and Bandaranzali coast were 10 and 10.71 alleles, respectively. The numbers of effective alleles were 7.05 and 7.74 alleles in each population.Allele frequency was found to have declined in wild fish due to inbreeding and genetic drift. The mean of observed heterozygosity values were 0.66 and 0.70 in Chamkhale and Bandaranzali coast, respectively.Approximately all of loci showed deviation from Hardy-Weinberg equilibrium. The genetic similarity and distance between the two populations were 0.316 and 0.684, respectively.The Fst value was 0.024 that indicates the low genetic differentiation between the two locations which could be explained by the low number of alleles in two populations. Also, the Natural Migration (Nm) between two stations was obtained 16.30. According to the analysis, it seems that Abramis brama hasn᾽t a desirable genetic diversity in the investigated regions.
 
Article
The prevalence of type 2 diabetes mellitus (T2DM) is rising dramatically in the Middle East, especially in the Islamic Republic of Iran, but the genetic basis of type 2 diabetes in Iran is poorly understood. Polymorphisms of hepatocyte nuclear factor-1α (HNF-1α) and glucagon-like peptide-1 receptor (GLP-1R) genes showed association with type 2 diabetes in several ethnic groups. In this study, we evaluated whether these markers confer susceptibility to T2DM in a diabetic population living in Mashhad (northeast of Iran). Genotyping of Ala98Val (HNF-1α) and Thr149Met (GLP-1R) was done by the restriction fragment length polymorphism-PCR (RFLP-PCR) method in the following groups: 1) early-onset diabetes (age at onset ≤ 35 years); 2) late-onset diabetes (age at onset > 35 years); and 3) control. Our results showed that CT (Ala/Val) genotype of HNF-1α was higher in the early-onset type 2 diabetic group compared to the controls but difference was not significant. We did not find the GLP-1R Thr149Met mutation in all participants. The prevalence of the HNF-1α (Ala98Val) and (GLP-1R) Thr149Met mutations has not been previously reported in Iranian participants. We conclude that these mutations are not a common cause of T2DM in our studied population.
 
Figure2. PCR analysis for detection of VP1 gene in transformed leaves of alfalfa. 1) 1 kb ladder; 2) plasmid pBI121VP1 (positive control); 3) transformed alfalfa plant; 4) wild type plant (negative control)
Quantitative measurement of VP1 gene transcription in transformed leaves of alfalfa via Real Time PCR. Data presented in this graph are obtained from three samples of transformed plants. 
Figure4. Dot blot assay for detection of recombinant protein in transiently transformed leaves of alfalfa. (1): protein sample of transformed leaves, (2) pure VP1-129-169 peptide as positive control, (3): protein sample of wild type plant (negative control)
Article
An Agrobacterium-mediated transient gene expression assay was carried out in alfalfa (Medicago sativa) leaves for expression of a chimeric gene encoding a part of capsid protein of Foot and Mouth Disease virus called VP1. The plant leaves were transformed via agroinfiltration procedure. The presence of the foreign gene and its expression in transformed plants were evaluated by polymerase chain reaction (PCR), real time PCR, protein Dot blot and ELISA. Moreover, gene expression in the transformed leaves was quantified by ELISA method. The results obtained in this investigation indicated high level of gene expression in alfalfa leaves, showing that transient gene expression can be applied as an effective and time-saving procedure for the production of recombinant proteins. The procedures for transformation, detection of recombinant protein and its application for molecular experiments are described in the study.
 
A) Down-regulation of miR-21 expression in HEK-293T stable cell lines exogenously expressing miRZip-21, in comparison to untransfected cells (p <0.0001). B) miR-21 expression level in U87-MG cells, 24 and 48 hours following co-culturing with HEK-293T cells stably expressing miRZip-21. As demonstrated no significant downregulation observed for treated cells.
miR-21 expression levels in exosomes isolated from miRZip-21 expressing HEK-293T cells, in comparison to the exosomes extracted from untransfected cells.
A) Expression of miR-21 in U87-MG cells exposed to the engineered exosomes isolated from conditioned media of miRZip-21 expressing HEK-293T cells by ultracentrifugation. B, C) Expression of PDCD4 and RECK in U87-MG cells following treatment with exosomes obtained from conditioned media of genetically modified HEK293T cells. As demonstrated similar expression pattern observed for miR-21 target genes, PDCD4 and RECK, 24 and 48 hours post-treatments. It should be notified that these modifications were not statistically significant.
Article
There are different subtypes of brain tumors, classified according to the origin of the abnormally proliferated glial cells. Glioblastoma multiforma (GBM) is the grade 4 of brain tumors, gliomas, with the least life expectancy. microRNAs (miRNAs) are small, single stranded, non-coding RNAs with 20-25 nt length with post-transcriptional gene regulatory activities. An altered expression of miRNAs is linked to developmental disorders and some diseases, most importantly cancers. miR-21 is a well-known microRNA, overexpressed in almost all cancer types, including brain tumors. It targets several genes with vital roles in cellular pathways involve in proliferation, invasion and metastatic behaviors. Exosomes are 30-100 nm extracellular vesicles which are packed with various molecules, including miRNAs. Here, we suppressed miR-21 expression level in HEK-293T cells by transfecting them with the miRZip-21 vector. However, when U87-MG cells were cultured in the presence of exosomes isolated from conditioned medium of engineered HEK-293T cells derived exosomes, we did not observe any suppressing effect on host cells' miR-21 expression level. Moreover, by analyzing the effects of miRZip-21-enriched cell's conditioned media on three other brain cell lines including 1321N1, A-172 and DAOY, cell type-specific effects of exocrine miRZip-21 were revealed. These data suggested that cell lines from different brain tumor subtypes could exert different responses to microRNA-based therapies, based on their cellular origin and clinical behaviors.
 
A schematic diagram of in situ hybridization procedure with LNA probe. 
Probe sequences and hybridization temperatures for each gene.
Nuclear signals of U6 snRNA; A) in NT2 cell line, B) in seminoma carcinoma FFPE tissue. 
A) MiR-21 cytoplasmic signals in FFPE tissue of seminoma carcinoma, B) Negative control without probe treatment. 
Cytoplasmic signals of miR-302, A) without counterstain, B) Negative control, C) slides were counterstained with nuclear fast red, blue signals show miR-302 localization, D) Negative control which is counterstained with nuclear fast red. There is no blue cytoplasmic signal in negative controls without probe treatment.
Article
MicroRNAs (miRNAs) are a group of short non-coding RNAs implicated in numerous fundamental cellular processes, and their disregulations have been linked to several pathologic conditions, mainly cancers. Determining tissue distribution of miRNAs is a prerequisite for understanding their exact functions during development, tissue homeostasis and abnormality. In situ hybridization is a powerful technique to delineate the sub-cellular localization and tissue distribution patterns of mRNAs as well as miRNAs. Due to the important role of miRNAs in tumorigenesis, we optimized an ISH technique for detection of two well-known miRNAs (miR-302 and miR-21) in formalin-fixed paraffin-embedded (FFPE) tumor samples along with a pluripotent embryonal carcinoma cell line, NTERA-2 (NT2). After fixation of cells on slides/sectioning of FFPE blocks, proteinase K digestion, probe concentration, antibody development and light sensitive color reaction were optimized for both the FFPE samples and cell line. Signals for U6 snRNA, as an internal control, were detected in the nuclei of the cells. MiR-21 and miR-302 expression was detected in the cytoplasm of FFPE samples of seminoma carcinoma and in NT2 cell line, respectively. In this study, we optimized ISH for miRNA detection in FFPE samples and NT2 cell line.
 
Restriction analysis of blue/white colonies isolated plasmids: lane 1 Eco RI/ Hind III lambda DNA marker, lane 2 non-digested plasmid from white colonies, lane 3 Xba I/ Bam HI digested plasmid isolated from white colonies which contain DREB1A fragment (circle), lane 4 none digested plasmid from blue colonies. 
Agarose gel electrophoresis representing PCR products for confirmation of success rate of DREB1A gene cloning. A, lane 1 to 9 show colony-PCR products separation, Lane 3 has a band corresponding to DREB1A fragment, lane 10 show positive control with genomic DNA of Arabidopsis thaliana . B, lane 1 lambda DNA marker, lane 2 negative control with ddH 2 O, lane 3 negative control with empty pBI121 vector, lane 4 colony-PCR containing pBI121::DREB1A , lane 5 positive control with genomic DNA of A.thaliana. C, lane 1 marker, lane 2 negative control with ddH 2 O, lane 3 positive control with genomic DNA of A. thaliana, lane 4 to 9 colony-PCR from sub-cultured 
Article
Environmental stresses affect plant growth and cause losses worth hundreds of million dollars to agricultural industry each year. Many genes are induced in response to environmental stresses. The DREB1A gene is a stress-inducible transcription factor which its ectopic over-expression improves plant tolerance to environmental stresses. To produce environmental stress tolerant plants carrying the DREB1A gene, the full length cDNA of the DREB1A gene was amplified from Arabidopsis thaliana Col-0 plants by gene specific primers and cloned into pGEMT-Easy vector, and transformed into E.coli. Presence of the DREB1A gene was confirmed by restriction analysis as well as DNA sequencing. A 668-bp XbaI/BamHI digested fragment of DREB1A gene from the pGEMT::DREB1A construct was sub-cloned into the pBI121 binary vector. The recombinant plasmids were transferred into Agrobacterium tumefaciens cells (strain LBA4404) and screened on LB medium supplied with kanamycin/rifampicin (50 mg/l). Positive bacterial colonies were selected based on colony-PCR analysis and saved for further application in plant materials.
 
Total testosterone concentration (a); expression of CX-43 mRNA (b), miR-1 expression (c), and miR-206 expression (d) in the left ventricular tissue of rats. NC group, normal control rats; ORX-S group, Castrated rats; ORX-T group, castrated rats that received testosterone enanthate 25 mg/kg/day. Data are expressed as mean ± SEM; Oneway ANOVA followed by Tukey's post hoc test. ***: P < 0.001;**: P < 0.01; *: P < 0.05. Abbreviations: CX-43, Connexin-43; miR-1, miRNA-1; miR-206, miRNA-206.
Article
Connexin-43 (Cx-43) plays axial roles in the propagation of action potentials and contractile coupling in the heart. Down-regulation of Cx-43 in the heart is associated with arrhythmia, dilated cardiomyopathy, and heart failure. To date, no studies have examined the effects of androgen deprivation therapy (ADT)-induced hypogonadism on the expression of Cx-43 in the heart. This study investigated the effects of testosterone deprivation and its replacement with testosterone enanthate on the expression of Cx-43 mRNA and the muscle-specific miRNAs miR-206 and miR-1, as two potential regulators of the Cx-43 protein expression in the ventricular tissue. Accordingly, 21 male Wistar rats were divided into three groups: Ι) Normal control, П) ORX-S: castrated rats serving as animal models for ADT and receiving the sesame oil as a solvent of testosterone enanthate for ten weeks, and Ш) ORX-T: these animals were castrated, receiving testosterone enanthate (25 mg/kg) for ten weeks. The relative expression of Cx-43 mRNA, miR-206, and miR-1 was determined by qRT-PCR. Cx-43 mRNA was found to be decreased in the ORX-S group. The Cx-43 mRNA was up-regulated after the administration of testosterone enanthate. There were no significant changes in miR-206, and miR-1 levels in the ORX-S and ORX-T groups compared to the controls. Our results indicated that testosterone should be regarded as an important factor in the regulation of Cx-43 mRNA expression in the heart, and testosterone deprivation may down-regulate the Cx-43 mRNA expression; however, it doesn't alter miR-1 and miR-206 levels. These results suggest that ADT-induced hypogonadism may put males at risk for cardiac dysfunctions.
 
Article
Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cell proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGC) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs was isolated from fat tissues and confirmed by their surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-pax6- pur lentiviral vectors in fibronectin supplemented medium, cells gradually showed the characteristic morphology of neuronal cells. QRT- PCR and ICC confirmed deriving of mainly RGC and marginally bipolar cells. The current investigation demonstrates the feasibility of differentiation of RGCs and bipolar cells from hADSCs using expression of PAX6 (+5a) in the medium supplemented by fibronectin.
 
PCR products were digested by BstUI restriction enzyme. The Arg allele BstUI digestion showed 169 and 127 bp fragments (lane 3). The Pro allele was not cleaved by BstUI, resulted in a single 296-bp band (lanes 1, 4, 5). The heterozygotes showed three bands (296, 169 and 127 bp) on gel electrophoresis (lane 2); Lane M: 50 bp DNA size marker. 
Article
Single nucleotide polymorphism in codon 72 of p53 gene (Arg/Pro) changes p53 protein structure and affects its activities. Hepatitis C virus (HCV) is believed to induce hepatocellular carcinoma and P53 polymorphisms have been associated with human cancers. The aim of this study was to evaluate genetic variants of codon 72 of p53 gene polymorphism in HCV patients and its relationship with HCV infection. The study was conducted on 67 HCV patients, who were referred to medical centers of Mashhad city, Iran, and 73 healthy people from the same region. Genotyping of codon 72 of p53 gene was performed by PCR-RFLP method. The distributions of different alleles of p53 polymorphisms did not differ significantly between groups. The respective proportions of Proline homozygotes, heterozygotes, and Arginine homozygotes were 37.31%, 35.82%, 26.86% in patients and 39.72%, 27.39%, and 32.87% in the control group respectively. However, we found no significant differenece for the allelic or genotype distribution between cases and controls. Our results indicated no strong evidence of association of the p53 polymorphism with HCV infection; however, further investigation is needed in different ethnic groups to elucidate the role of this polymorphism in HCV infection.
 
Schematic representation of human ABC transporters existence and their role in tissue defense and organ regeneration by excretion, absorption and distribution of toxic substances within/out of the cells.
Two primary types of membrane associated ABC transporter proteins [A] Full transporter shows mirror image halves separated by a flexible linker region essential for catalytic activity. Each half of the molecule is a 6-transmembrane spanning domain (6TM) that blocks the transporter proteins to the plasma membrane [B] Half transporters are consist of only one halve and require membrane association with other half transporter to form the pure complex. There is no linker region present in the half transporter proteins. 
Role of ABC transporters in NSCs proliferation and differentiation. Usually during NSCs proliferation expression of ABC transporters increases and decreases during differentiation 
Article
Increasing incidence of central nervous system (CNS) disorders has become a major challenge for both basic and clinical scientific society to develop novel therapeutic models for treatment. The knowledge of stem cells has added a new dimension in the research towards finding more appropriate targets responsible for the disease fate determination. As stem cell research is progressing day by day in routine research laboratories there is great hope to find suitable therapeutic targets for complete cure of the CNS disorders. Discovery of ABC transporters in animal tissues has emerged as new spot for several disease prognosis and therapeutic target. ABC transporters are membrane proteins expressed in various organs like liver, kidney, blood-brain barrier, blood-testis barrier etc. It is involved in various important cellular processes such as absorption, distribution and excretion of drugs, xenobiotics and endogenous compounds showing their role in tissue defense and organ regeneration. The current review explains about the role of ABC transporters in CNS pathogenesis and defense adding stem cells therapeutic strategies.
 
Schematic procedure of TA cloning used in this study for constructing the reference standard. 
Agarose gel electrophoresis of PCR products obtained under optimized conditions for detection of B. subtilis (212 bp). Extracted DNA from cultured cells of B. subtilis in nutrient broth as positive control (lane 1), 100 bp DNA marker (lane 2), extracted DNA from inoculated dough with B. subtilis (lane 3), extracted DNA from dough artificially contaminated with B. amyloliquefaciens, B. cereus, B. licheniformis, Bacillus spp., S. aureus, E. coli and Lactobacillus spp., representing the negative controls with ca. 105 CFU/g of each bacterium (lanes 4 to 10, respectively).
(a) Real time PCR amplification of B. subtilis aprE gene. (b) Melting curve analysis of SYBR green real time PCR product of B. subtilis (in broth culture as positive control and induced contaminated dough) after 40 cycles in presence of negative control bacterial species used for conventional PCR. The melting temperature is calculated as 82.2 oC. No nonspecific peaks are present.
Article
In this present study, a SYBR green based real time PCR assay has been developed for specific detection and quantification of Bacillus subtilis in dough used for bread making. New primer pairs were designed to amplify a 212 base pair fragment of the aprE gene. Specificity of these primer pairs was confirmed with conventional and real time PCR methods. Standard curves constructed using the threshold cycle (CT) versus copy numbers of B. subtilis showed good linearity for reference standards of cloned insert (R 2 =0.999, slope=-3.035) and also induced contaminated dough (R 2 =0.988, slope=-3.142), and the melting temperature (Tm=82.2 o C) was consistently specific for the amplicon. Limits of detection were 200 and 2000 colony forming units (CFUs) per ml or g of these samples, respectively. This real time PCR offers a fast tool with high sensitivity and specificity for detection and quantification of this rope-forming pathogen in dough used for bread making.
 
Article
Epithelial ovarian cancer (EOC), as a challenging disease among women with poor prognosis and unclear molecular pathogenesis, each year is responsible for 140000 deaths globally. Recent progress in the field revealed the importance of proteins as key players of different biological events. Considering the complicated protein interactions, taking a deeper look at protein-protein interactions (PPIs) could be considered as a superior strategy to unravel complex mechanisms encountered with regulatory cell signaling pathways of ovarian cancer. Hence, PPI network analysis was performed on differentially expressed genes (DEGs) of ovarian cancer to discover hub genes which have the potential to be introduced as biomarkers with clinical utility. A PPI network with 600 DEGs was constructed. Network topology analysis determined UBC, FN1, SPP1, ACTB, GAPDH, JUN, and RPL13A, with the highest Degree (K) and betweenness centrality (BC), as shortcuts of the network. KEGG pathway analysis showed that these genes are commonly enriched in ribosome and ECM-receptor interaction pathways. These pivotal hub genes, mainly UBC, FN1, RPL13A, SPP1, and JUN have been reported previously as potential prognostic biomarkers of different types of cancer. However, further experimental molecular studies and computational processes are required to confirm the function and association of the identified hub genes with epithelial ovarian cancer prognosis.
 
Mean of concentration±SD for different elements in the liver and kidney of the sea water fish and lagoon fish of the Iranian Caspian Sea (*p <0.05).
Characteristic load for PC1, PC2 and PC3 obtained by principal component (PCA) analysis for elemental concentrations of the fish liver and kidney between habitats (sea water and lagoon) in the water of Iranian Caspian Sea
Article
The Caspian Sea is the largest inland body of water in the world and so has both common characteristics of seas and lakes with over 153 fish species which inhabit the sea and its basin. However, little is known about the trace element (TE) contaminations (TECs) in its tissues. In the present study, 122 specimens of three fish species including Rutilus caspius (Roach, n=71), Leuciscus aspius (Asp, n=20), and Tinca tinca (Tench, n=31) were collected from three different fisheries regions (i.e. Astara, Anzali and Kiashahr) of the southern part of the Caspian Sea from September 2017 to June 2018. Inductively coupled plasma optical emission spectrometry (ICP-OES) was employed to measure TE levels in different fish tissues. An attempt was made to assess possible influences of habitat on element accumulation in the liver and kidney of three fish species in the southwest of the Caspian Sea basin. Some elements including Ca, K, Mg, P, S, Sc, and Sr showed different concentrations in the liver and kidney. Also their levels were significantly different between freshwater resident (Tench) and marine (Roach) species (p < 0.05). The differences among TECs in the liver and kidney of Roach, Asp and Tench were reduced to three components using principal component analysis (PCA). Results indicated that 83.60% of the total variability is related to TEs such as Cu, Fe, Sr, Ca, S, Na, Mg, K, and Al. The impact of habitat variability on the element accumulation was confirmed through linear chart obtained for liver and kidney (as body filtering organs) of Roach and Asp as marine residents as well as Tench as a freshwater resident. This could illustrate the borderline created by these habitats Element accumulations in liver and kidney tissues of some bony fish species during inhabiting salt and fresh water in the southwest of the Caspian Sea.
 
Mean of concentration±SD for different elements in the liver and kidney of the sea water fish and lagoon fish of the Iranian Caspian Sea (*p <0.05).
Characteristic load for PC1, PC2 and PC3 obtained by principal component (PCA) analysis for elemental concentrations of the fish liver and kidney between habitats (sea water and lagoon) in the water of Iranian Caspian Sea
Article
The Caspian Sea is the largest inland body of water in the world and so has both common characteristics of seas and lakes with over 153 fish species which inhabit the sea and its basin. However, little is known about the trace element (TE) contaminations (TECs) in its tissues. In the present study, 122 specimens of three fish species including Rutilus caspius (Roach, n=71), Leuciscus aspius (Asp, n=20), and Tinca tinca (Tench, n=31) were collected from three different fisheries regions (i.e. Astara, Anzali and Kiashahr) of the southern part of the Caspian
 
Light micrographs of mitotic metaphase chromosomes prepared from different populations of Achillea tenuifolia. A, Population No. 1 with chromosome number 2n=2x=18; B, Population No. 2 with chromosome number 2n=4x=36. C-P, The populations 3-9 with two different chromosome numbers 2n=2x=18 and 2n=4x=36. Arrow indicated B-chromosomes or satelites chromosomes in the figures. Bar in all figures= 5μm.
Light micrographs of mitotic metaphase chromosomes prepared from different populations of Achillea biebersteinii. A, Population No. 1 with chromosome number 2n=2x=18; B, Population No. 2 with chromosome number 2n=2x=18. C-N, The populations 3-8 with two different chromosome numbers 2n=2x=18 and 2n=4x=36. Arrow indicated B-chromosomes or satelites chromosomes in the figures. Bar in all figures= 5μm.  
Article
Chromosome counting was performed in nine populations of Achillea tenuifolia Lam and eight populations of A. bieberestinii Afan (Asteraceae) collected from Hamedan and Kermanshah provinces in the west of Iran. Chromosome numbers in both species varied from 2n=2x=18 to 2n=4x=36. Some populations of both species showed (2n=4x=36) chromosome number that is the first report as polyploidy levels. Aneuploidy is also the first report for both species. Diploid and tetraploid individuals were observed in some populations at the same locality. B-chromosomes were observed in some populations of both species. The results indicated that polyploidy is a common future in this species similar to other Asteraceae plants.
 
The most important Amino Acid Composition selected by different attribute weighting algorithms in discriminating allergen proteins from non-allergen proteins. 
Article
Allergens are proteins or glycoproteins which make widespread disorders that can lead to a systemic anaphylactic shock and even death within a short period of time. Understanding the protein features that are involved in allergenicity is important in developing future treatments as well as engineering proteins in genetic transformation projects. A big dataset of 1439 protein features from 761 plant allergens and 7815 non-allergen proteins was constructed. Thereafter, 10 different attribute weighting algorithms were utilized to find the key characteristics differentiating allergens and non-allergen proteins. The frequency of Leu, Arg and Gln selected by different attribute weighting algorithms with more than 50% confidence, including attribute weighting by Weight_Info Gain, Weight Chi Squared, Weight_Gini Index and Weight Relief. High amount of Gln and low percentage of Leu and Arg discriminate plant allergens from non-allergens
 
Schematic structural representation of retinoic acid isoforms including all-trans (At), 9-cis (9c), and 13-cis (13c) retinoic acids (RAs). At-RA is the main active isomer of RA (http://pubchem.ncbi.nlm.nih.gov/).
Docking energy amounts are shown for various complexes of At-RA, 9c-RA, and 13c-RA isoforms with different VEGF receptors, including VEGFR-1 D3 (5T89), VEGFR-2 D3 (2X1X), and VEGFR-2 D3 (2X1X).
Energy minimization (first and end energy) for VEGFRs.
Article
In this study, putative interactions between all of the retinoic acid (RA) ligands (all-trans (At), 9-cis (9c), and 13-cis (13c)), and VEGF receptors (VEGFR-1, -2 and -3) were investigated. It was performed considering the glycosylation status of the receptors to achieve a more reliable mode of interactions based on glycomics. We found that RAs may have a higher affinity for ligand-binding domains in VEGFRs. Furthermore, all RA isomers can strongly attach to VEGFR-3 receptor in comparison to other ones. It was also demonstrated that receptor dimerization of RAs may be less targeted. Moreover, regarding post-translational modifications, glycosylated structures showed conflicting binding energies. RAs may target the human vasculature, specifically lymph vessels, through VEGFR-3. In addition, the ligand binding-mediated activation of VEGFRs may be affected by these agents. Also, the glycosylation status of the receptors can interfere with these manners. Furthermore, our results confirmed that the consideration of carbohydrates in crystal structures is essential for a better interpretation of ligand/receptor interactions during drug discovery studies. Even though these observations improved our understanding of the binding patterns of RAs to VEGFRs, validation of these results needs further analysis to introduce these biomolecules as anti-VEGF remedies.
 
Human cultured RPE cells grew as a monolayer of cuboidal epithelial cells arranged in a regular hexagonal pattern (magnification: (A) 10x40, (B) 10x20). 
Primer Sequences
Human cultured RPE cells subjected to ICC with cytokeratin 8/18 and RPE65 antibodies. (A) Nuclei stained blue with DAPI. (B) The RPE cells stained positively for the FITC-conjugated cytokeratin 8/18 antibody (green). (C) Merged image (FITC-labeled cytokeratin 8/18 and DAPI) (magnification: 10x40). (D) DAPI-stained RPE cell nuclei (blue). (E) The RPE cells stained positively for the RPE65 antibody (green). (F) Merged image (FITC-labeled RPE65 and DAPI) (magnification: 10x20). 
Quantitative real-time RT-PCR analysis of MITF-A, MITF-H, OTX2, TYR and DCT in passages 1-7 of cultured neonate RPE cells. RPE cell preparation and RNA extraction were performed as described in methods. Relative gene expression was determined by quantitative real-time PCR. mRNA levels were normalized to GAPDH and presented as percentages of control values. (B) MITF-H at passage 7, (C) OTX2 at passages 4-7, (D) TYR at passages 4-7 and (E) DCT at passages 2-7 decreased compared to the other passages specially regarding passage 1. *P <0.05
Article
Retinal pigment epithelium is responsible for maintaining the structural integrity of the retina by an efficient defense against free radicals, photo-oxidative exposure and light energy. For this purpose the main RPE line of defense is melanosomes. Melanin content of retinal pigment epithelium cells in adults and neonates reveals remarkable variations. In the current study we compared melanogenic factors gene expression levels in human adult and neonate RPE cells in culture. RPE cells from adult and neonate human eye globes were isolated and then cultured in DMEM: F12 (1:1) containing 10% FBS. Expression of RPE65 and Cytokeratin 8/18 markers in isolated cells was confirmed by immunocytochemistry. To evaluate the responsible factors in the pathway of melanin biosynthesis, gene expression of orthodenticle homeobox 2, microphthalmia-associated transcription factor A and H as prominent transcription factors, tyrosinase and dopachrome tautomerase as effective enzymes in the melanin biosynthetic pathway were examined in human neonate derived RPE cells compared to human adult derived RPE cells in culture. According to the Real-Time RT-PCR data, gene expression of MITF-H, OTX2, TYR and DCT in RPE cells of the neonates showed a significant increase compared to the adults. With increasing passage number, gene expression of MITF-H, OTX2, TYR and DCT showed remarkable decline. According to the role of OTX2 and MITF-H as the main transcription factor effectors on the TYR and DCT, restoration of OTX2 and MITF-H gene expression may be retain melanin content in RPE cells.
 
RAD52 substitution by KANMX cassette in deletion mutants (No.1-5) of yeast haploid strains of BY4742 (No. 1 and 2) and of BY4741 (No. 3,). Bands in lanes 1, 2, 3 obtained from primer combinations P5 and P6 (635 bp), and in lanes 1', 2', 3' obtained from primers combinations P7 and P8 (560 bp). M: DNA molecular ladder of 10 kb.
Primers designed and used in this study.
Presence of both MATa and MATα loci in yeast rad52 diploid deletion mutants constructed by mating haploid strains of BY4742 and BY4741. Lane 1: MATa in haploid strain BY4741 (492 bp); lane 2: MATα in haploid strain BY4742 (369 bp); lane 3: MATa and MATα in diploid strain BY4743; lanes 4, 5, 6, 7: MATa and MATα in rad52 diploid deletion strains in yeast BY4743 background. M: DNA molecular ladder of 10 kb.
Article
Agrobacterium tumefaciens is capable to transfer genes across kingdoms. It can genetically transform not only plant cells, but also many other bacterial, algal, fungal, animal and human cells. This depends on the interactions among a variety of both Agrobacterium and host genes. Inside the host cell, RAD52 which is involved in DNA repair is a key gene determining integration of T-DNA by homologous recombination. Here, using Saccharomyces cerevisiae haploid strains BY4741 and BY4742, a rad52 diploid deletion strain was constructed in yeast BY4743 background. This model organism was employed to show that RAD52 deletion severely decreases frequencies of Agrobacterium genetic transformation mediated by either an integrative T-DNA or a circular non-integrative T-DNA. Indeed, the frequencies of such Agrobacterium-mediated transformation (AMT) decreased by ca. 25-fold, compared to wildtype BY4743. Hence, host RAD52 deletion might affect AMT by a mechanism which differs from its only involvement in DNA repair in yeast.
 
PCR amplification of hph selection marker gene (544-bp) in mitotically stable transformants (No.5- 14) of C. gloeosporioides obtained by A. tumefaciens strain pSDM2315 (lanes:5-9), and by A. tumefaciens strain pBSY90 (lanes:10-14). Lanes 2 and 3 include positive controls (from binary vectors pTAS10, and pBin- GFP-hph ). Lane 4 represents negative control. DNA ladder: 1000 bp ladder (Cinnagene). The observed PCR bands accord to 544 bp, as expected. 
Gfp expression in a representative hygromycin-resistant transformant's hyphae of C. gloeosporioides, after ATMT with pBSY90.
Different range growth and morphology/pigmentation of six C. gloesporioides transformants (A-F) as compared to their parental wild type (G) on PDA plates 12 days after incubation at 25 o C
Article
The fungus Colletotrichum gloeosporioides is the causative agent of anthracnose disease of many tropical, subtropical and temperate fruits, and a microbial source of the anticancer drug, Taxol. Here, we introduce an optimized Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for genetic manipulation of this fungus using hph and gfp-tagged hph genes as selection markers. Results showed that falcate spores can be easily used instead of protoplasts for transformation. Several experimental parameters were shown to affect transformation efficiencies, among which the length of co-cultivation, the ratio of fungal conidia to bacterium during co-cultivation, the kind of membrane during co-cultivation, and the kind of fungal growth medium during transformants selection, showed the highest influences on ATMT frequencies. Results indicated that the optimal ATMT of C. gloeosporioides was achived after 3 days of co-cultivation, at 107 per ml fungal conidia, via the use of Fabriano 808 filter paper and Czapek's culture medium. Successive subculturing of transformants on selective and non-selective media demonstrated the stable expression of transgens, and subsequent PCR based analyses of transformants revealed the presence (100%) of transferred genes. Flourescence microscopy analyses showed a punctuate pattern for localization of an expressed Gfp-tagged Hph protein inside fungal hyphae. The optimized ATMT protocol generated mutants that showed different phenotypes based on their vegetation and pigmentation. This suggests the possible applicability of this technique for functional genetics studies in C. gloeosporioides, through insertional mutagenesis.
 
Figure2. PCR analysis for detection of VP1 gene in transformed leaves of alfalfa. 1) 1 kb ladder; 2) plasmid pBI121VP1 (positive control); 3) transformed alfalfa plant; 4) wild type plant (negative control) 
Quantitative measurement of VP1 gene transcription in transformed leaves of alfalfa via Real Time PCR. Data presented in this graph are obtained from three samples of transformed plants. 
Figure4. Dot blot assay for detection of recombinant protein in transiently transformed leaves of alfalfa. (1): protein sample of transformed leaves, (2) pure VP1-129169 peptide as positive control, (3): protein sample of wild type plant (negative control) 
Figure5. Quantification of recombinant VP1 expression in three transgenic alfalfa plants by ELISA. 
Standard curve of VP1 protein 
Article
An Agrobacterium-mediated transient gene expression assay was carried out in alfalfa (Medicago sativa) leaves for expression of a chimeric gene encoding a part of capsid protein of Foot and Mouth Disease virus called VP1. The plant leaves were transformed via agroinfiltration procedure. The presence of the foreign gene and its expression in transformed plants were evaluated by polymerase chain reaction (PCR), real time PCR, protein Dot blot and ELISA. Moreover, gene expression in the transformed leaves was quantified by ELISA method. The results obtained in this investigation indicated high level of gene expression in alfalfa leaves, showing that transient gene expression can be applied as an effective and time-saving procedure for the production of recombinant proteins. The procedures for transformation, detection of recombinant protein and its application for molecular experiments are described in the study.
 
Article
Aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor and its induction may result in suppressing of cell proliferation in colorectal cancer (CRC). Cucurbitacin D (CucD), E (CucE) and I (CucI) are plant derived metabolites that inhibit cancer cells. This study aimed to evaluate the possible potency of the cucurbitacins for activation of AHR expression in CRC cell lines SW-480 and HT-29. The MTT assay was used to find the IC50 value of the metabolites in the cell lines. Afterwards, the cells were incubated with the IC50 concentrations of the cucurbitacins and AHR-mRNA expression assessed using RT-PCR. The IC50 values of CucD, CucE, and CucI were 4.5, 6.8, and 3.8 μM in HT-29 cell line and 35, 19, 17.5 μM in SW-480 cells, respectively. The SW-480 cells were more resistant against cucurbitacins in comparison with HT-29 cells and all three cucurbitacins led to more AHR-mRNA expression in HT-29 cells. CucE had the lowest effect on AHR-mRNA expression in the cell lines and CucI was a common metabolite for both HT-29 and SW-480 cells, which showed the lowest IC50 value (the highest toxicity) and the highest effect on AHR-mRNA expression. CucI may have a potential AHR-induction role and it could be applicable as an AHR-expression inducer in CRC studies.
 
Schematic image of alternative splicing of lncRNA
The results for Quality control trimming and mapping had gained per 12 samples
Article
Long non-coding RNAs (lncRNAs) compose a plentiful category of transcripts that have gained increasing importance because of their roles in different biological processes. Although the function of most lncRNAs remains unclear. They are implicated in epigenetic regulation of gene expression, including muscle development and differentiation. We aimed to identify the effect of novel lncRNAs (Alternatively spliced) and their target genes on two stages of sheep skeletal muscle growth and development. FastQC files have been used to examine the quality control and the Trimmomatic program for trimming low-quality reads from twelve longissimus dorsal muscle tissue samples (including six young and six old from Texel sheep). Hisat2, Cufflink, Cuffmerge, and Cuffdiff investigated the expression levels. Novel lncRNAs (Alternative spliced) were distinguished using NONCODE databases and Cuffcompare software. In addition, the lncRNA-mRNA interactions and regulatory network visualization were identified via RIsearch and Cytoscape software, respectively. Those 139 novel lncRNA (Alternative spliced) transcripts had been recognized, probably 65 lncRNAs interacted with their target genes and regulated sheep skeletal muscle growth and development. Three novel lncRNA transcripts (TCONS_00041386, TCONS_00050059, and TCONS_00056428) showed a strong association and five transcripts (TCONS_00055761, TCONS_00055762, TCONS_00055763, TCONS_00055764, and TCONS_00055770) had made complex network correlations with mRNAs. Our research provided more knowledge of the associated mechanisms with novel lncRNAs, which could play a role in regulating sheep skeletal muscle tissue development and growth.
 
Article
Recent genome-wide association studies have introduced several genetic variants which contribute to the late-onset Alzheimer's disease (LOAD). Polymorphisms of CHAT, TOMM40, and SORL1 genes have been reported to be associated with the LOAD phenotype. This study was endeavored to evaluate the association of the CHAT rs3810950, TOMM40 rs1160985 and SORL1 rs11218304 polymorphisms with the LOAD in the Turkish-speaking Azeri population of northwest Iran. In a case-control study, we included 174 cases: 88 cases with LOAD diagnosis and 86 healthy individuals. Peripheral blood samples were collected and the genomic DNA of all participants were extracted. Genotyping was carried out by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. We did not observe any significant association between the CHAT rs3810950 and SORL1 rs11218304 alleles with the LOAD. However, both the TOMM40 rs1160985 minor allele T and TT genotype showed significant negative associations with the LOAD. Hence, the TOMM40 rs1160985 polymorphism could be considered as a protective genetic factor against the LOAD in the Turkish-speaking Azeri population of northwest Iran.
 
Article
Background: Amniotic membrane derived stem cells (AMSCs) have considerable advantages to use in regenerative medicine and their anti- inflammatory effects, growth factor secretion and differentiation potential make them suitable candidates for stem cell therapy of nervous system. The developing and neonatal brain contains a spectrum of growth factors to direct development of endogenous and donor cells. Using an in vitro model system, we investigated the plasticity and potential of mouse AMSCs to differentiate into neural cells in response to neonatal mice brain extracted medium. Methods: Mouse amniotic membrane stem cells were isolated from embryos, cultured in presence of medium derived from neonatal mouse brain medium and immunohistochemistry and flow cytometry analyses were used to explore the neural differentiation of them. Results: Isolated amnion membrane stem cells showed high rate of viability and proliferation and presented neural characters in the presence of neonatal brain extracted medium such morphological changes and Nestin and Map-2 expression. Conclusion: In conclusion, results from this study showed that amnion membrane derives stem cells are potent stem cells to respond to environmental signals promoting them to neural fate.
 
Genomic DNA of about 150-400 ng extracted by CTAB in C. cyminum 
Comparison of genomic DNA template stability in two ecotypes of cumin seedlings exposed to different Cd concentration. 
Article
Cadmium, a metal widely used in industrial processes, has been recognized to be a highly toxic and dangerous environmental pollutant. Random amplified polymorphic DNA (RAPD) test is a feasible method to evaluate the toxicity of environmental pollutants on vegetal organisms. Herein, two Iranian ecotypes of Cuminum cyminum (cumin) plantlets following exposure to cadmium (Cd) concentrations of 300–1050 μM for 7 days were screened for DNA genetic alterations by DNA fingerprinting. 10 RAPD primers of 50–70% GC content were found to produce unique polymorphic band profiles on treating cumin seedlings. After Cd treatment, significant changes were observed in RAPD profiles of both ecotypes. These changes included variation in band intensity, disappearance of bands, and appearance of new PCR products in comparison to the control group, and they were dose dependent. These results indicated that genomic template stability (GTS, a qualitative measure reflecting changes in RAPD profiles) was significantly affected at the above Cd concentrations. The GTS index in both ecotypes gradually decreased with an increase in Cd concentrations. These findings suggest that DNA polymorphism detected by RAPD analysis could be a powerful eco-toxicological tool to evaluate the genotoxic effects of cadmium on plants.
 
Article
Long non-coding RNAs (lncRNAs) have recently found to have important regulatory roles, and their aberrant expressions and functions are directly linked to carcinogenesis. Both urinary bladder and breast tumors are prevalent neoplasms, with high rates of incidence. To identify a potential expression alteration of the recently discovered "anti-differentiation non-coding RNA, (ANCR), during tumorigenesis, we initially assessed its expression in several cancer cell lines (LNCAP, MCF-7, Ht-29, 5637, A549, HepG2, and PC3) and then compared its expression variability in tumor vs. non-tumor samples of bladder and breast. Here, ANCR expression profile was studied by qRT-PCR in paired tumor and marginal non-tumor samples obtained from patients that had been referred to the Labbafi-Nejad and Imam Khomeini Hospitals, respectively. Our data revealed a significant upregulation (p = 0.003) of ANCR in breast tumor tissues, in comparison to non-tumor marginal specimens from same patients. Similar upregulation was also detected in bladder tumor samples, however, this alteration was not statistically significant (p ≥ 0.05), probably due to small number of samples (n = 10). In conclusion, our results suggest a possible role of ANCR in tumorigenesis of bladder and breast tissues, as well as its potential usefulness as a novel diagnostic biomarker for bladder and breast tumors.
 
Article
Potato (Solanum tuberosum L.) auxiliary buds c.v. White Desiree were cultured in MS medium containing 0, 50, 100, 150 and 200 µM concentrations of silver thiosulfate (STS) under in vitro condition. After eight weeks, the effect of silver ions (Ag +) in the form of silver thiosulfate complex (STS), as an ethylene action inhibitor, on chlorophyll contents of leaves, ascorbate peroxidase, guaiacol peroxidase and catalase activities of roots and leaves were studied. Application of silver (STS) in culture medium increased chlorophyll content comparing to the control plants significantly. After treatments of potato plants with STS, ascorbate peroxidase and guaiacol peroxidase activities in roots were higher than shoots while catalase activity was higher in leaves than roots. However, increasing of STS concentration in the culture medium resulted in higher activities of antioxidant enzymes with some variations.
 
Chromosome measurements (in µm) and classification of Aphanius shirini chromosomes (Ch. No.: Chromosome number; LA: Long arm; SA: Short arm; TL: Total length; AR: Arm ratio; CT: Chromosome type; Sm: Submetacentric; St: Subtelocentric).
Article
The karyological and cytological characteristics of an endemic cyprinodont fish of Iran, Aphanius shirini have been investigated for the first time by examining metaphase chromosomes spreads obtained from gill epithelial and kidney cells. The diploid chromosome number of this species is 48. The karyotype consisted of one submetacentric and 23 subtelocentric pairs of chromosomes (2Sm + 46St). The chromosome arm number (NF) is 50. Sex chromosomes were cytologically indistinguishable in this tooth-carp. Based on the present and previous reported diploid chromosome number for other cyprinodont species, it can be suggested that the diploid chromosome number of 2n = 48 is the modal number of the cyprinodont fish.
 
Article
Green plants have emerged as ideal platforms for production of recombinant vaccine during recent decades. Various antigens relating to a large number of animal and human diseases have been studied in different plant species for production of recombinant vaccines. Despite the unique advantages of plant systems as green factories for production of recombinant vaccines, there are some major hurdles that have prevented commercial production of plant-based vaccines. In this review, theoretical background and practical applications of plant system for production of various recombinant vaccines are discussed.
 
Proline content in shoot (A) and roots (B) of transgenic and non transgenic tobacco seedlings. Values are means ± Sd. Uncommon letters are significant based on Tukey test (P < 0.05). 0, 5, 10, 20 30% of PEG are equal to 217, 264, 320, 637, 1292 mmol/kg respectively.  
Ascorbat peroxidase (APX) activity of leaves of tobacco seedlings in response to drought stress. Values are means ± SE. Uncommon letters are significant based on Tukey test (P < 0.05). 0, 5, 10, 20 30% of PEG are equal to 217, 264, 320, 637, 1292 mmol/kg respectively.  
Effect of PEG on MDA content in transgenic and non transgenic tobacco plants. Values are means ± SE. Similar letters indicate not significant based on Tukey test (P < 0.05). 0, 5, 10, 20 30% of PEG are equal to 217, 264, 320, 637, 1292 mmol/kg respectively.  
Article
In this study proline content and activity of catalase (CAT), and ascorbate peroxidase (APX) and level of lipid peroxidation in terms of malondialdehyde (MDA) content were measured in transgenic tobacco (Nicotiana tabacum cv. Wisconsin), over expressing a Δ-1-pyrroline-5-carboxylate synthase (P5CS) gene, and non transgenic plants as control. Drought stress was applied using polyethylene glycol (PEG) 6000 at concentrations of 217, 264, 320, 637, 1292 mmol/kg equal to (0, 5, 10, 20, 30% respectively). Proline content, especially in transgenic plants, was increased in leaves and roots significantly. CAT and APX activities increased under drought stress and the highest activity was observed in 10 and 20% of the PEG treatment. MDA content was increased by increasing of PEG and the highest MDA content was revealed in transgenic and non transgenic plants at 20% and 30%, respectively. Our results suggest that P5CS is an inducible gene and over production of proline and induction of CAT and APX activities are involved in drought tolerance mechanism.
 
Top-cited authors
Omid Dayani
  • Shahid Bahonar University of Kerman
Mohammad Reza Mohammadabadi
  • Shahid Bahonar University of Kerman
Masood Asadi Fozi
  • Shahid Bahonar University of Kerman
Amin Khezri
  • Shahid Bahonar University of Kerman
Moslem Shojaei
  • Ferdowsi University Of Mashhad