مجله زیست شناسی ایران
سال نوزدهم شماره 3 (پاییز 1385)

The species of Rhizoctonia solani is one of the most diverse and complex species that morphologically is similar to Basidiomycetous imperfect fungi. Its broad range and the lack of diagnostic characters create difficulties in the study of taxonomy and population genetics of this fungus. In this study zymogram of pectinase was used as a molecular marker to determine genetic variation of R. solani AG-2 which is known as the cause of brown patch of lawn disease. In this regard 252 out of 256 isolates recovered from lawn were identified as R. solani AG-2, based on the number of nuclei per cell and anastomosis reaction. In pectic zymogram analysis of the isolates tested two loci, one for polygalacturonase and the second for pectinestrase were identified. Based on polymorphisims in these two loci, ten zymogram patterns were distinguished for 252 isolates tested. It seems not only this technique is a fast and reliable method to identify anastomosis groups of R. solani but also it could employed to study intraspecific variation of this fungus.

Ubiquitin is a small protein of 76 amino acid residues that has attracted the concern of researchers of different science due to its multiple functions in the most biological process. It contributes in process such as cellular proteolysis, replication control and gene expression, maintenance of chromatin structure, cell differentiation, gametogenesis and etc., which shows numerous function of this vital protein. The immunochemical methods (ELISA, radioimmunoassay, Immunofluorecence) are sensitive and rapid techniques for the analysis of Ubiquitin and other such biological molecules which all use specific antibodies as detector. Therefore preparation of specific antibody directed against Ubiquitin is a valuable tool in detecting and assessing of this protein. In this study we attempted to purify Ubiquitin from human red blood cells and produce specific antiubiquitin antibody and conjugate these antibodies in order to employ in immunochemical studies. In this study Ubiquitin was first purified from packed blood cells by several steps including rapid denaturation of protein at 90°c, precipitation of Ubiquitin and other protein in 90% saturated ammonium sulfate and chromatography on DEAE-Sephadex. In the next step to increase the immunogenecity of Ubiquitin, cross linking of this protein to form polyubiquitin was attempted by employing glutaraldehyde. The prepared immunogenic polyubiquitin, were injected multiportally in to the rabbit. The reactive antibody was detected against polyubiquitin, using enzyme linked immunoassay. In order to isolate rabbit serum antibody, Protein A affinity chromatography was employed. Free Ubiquitin was coupled to CNBr-activated Sepharose 4B so that by passing rabbit antiserum over the column antiubiquitin antibody was purified. Specificity of antibodies was also examined using enzyme linked immunoassay. Finally antiubiquitin antibodies were conjugated with Flourscein Isothiocyanate. Results on SDS-Gel Electrophoresis of purified Ubiquitin and antiubiquitin antibody from Ion exchange column and affinity column appeared as a single band, indicating protein purity. It has also confirmed the specificity of purified antibodies referred to antiubiquitin antibodies by using ELISA. Use of such purification methods gave a yield of 1mg Ubiquitin from 100ml packed blood cell and 0/6 mg antiubiquitin antibodies from 8 ml immune serum. Study and identification of Ubiquitin function, play an important roles in biological research therefore preparation of antiubiquitin antibodies as an initial tool of these researches in immunochemistry studies is very important and there has been much interest in the production of these antibodies. The offered method in the preset study for purification of Ubiquitin and antiubiquitin antibodies is a reliable way to achieve this aim. Access to Antibodies conjugated with Fluorochrome molecules in this study; provide a tool in Ubiquitin studies by Immunofluorecence methods.

Streptomycin is an aminoglycosidic antibiotic that is produced by Streptomyces griseus. The genetics of streptomycin production is well known in Streptomyces griseus, for which more than 25 clustered genes have been described that encode biosynthesis, regulatory and transport functions. The main goal in this research was production of elevated levels of streptomycin, using transgenic Streptomyces. To gain this, the regulatory gene for streptomycin production (strR) was needed. In this research DNA was extracted and purified from Streptomyces griseus. Different sets of primer were designed for this research purpose and then the strR gene was amplified by PCR. Identify of PCR product of strR gene, was confirmed using gel electrophoresis, Nested-PCR and PCR-RFLP techniques.

The main active compounds of Lactogen herbal extracts are pectin and -glucan which are polysaccharide structure. Studies on the effect of these substances on hypophysis fragments indicated their potential to increase prolactin secretion. In the present study, we used GH3/B6 cell line which is known to secret prolactin and growth hormone. Western blotting following band densitometry via Total lab software showed prolactin amount in the presence of -glucan. In our study GH3/B6 cells were treated with various concentration of -glucan ranging from 50 to 300 gr/ml for a time period of 24 hours or 48 hours. -glucan effect on prolactin secretion was compared to control cells (in the absence of -glucan) and positive control cells treated with 50 nM of Thyrotropine Releasing Hormone (TRH). TRH is one of the best prolactin secretion stimulator in anterior hypophysis cells. These results indicate that 24 hours incubation of these cells in the presence of 50, 100, 200 gr/ml (p<0.01) and 300 gr/ml (p<0.05) of -glucan can significantly increase prolactin secretion in comparison with control. In 48 hours incubation with -glucan, prolactin secretion was significantly increased in 100, 300 (p<0.05) and 200 gr/ml (p<0.01) in comparison with the control. In addition, phase contrast microscopy showed increased appearance of secretory granules which could correlate with increased prolactin secretion.

Without any doubt, the issue of ER export is far from settled and discussions have been ongoing for almost two decades. Two models are currently competing for general acceptance. One model describes ER export as a non-selective diffusion into anterograde ER-derived transport vesicles (bulk-flow). The second model postulates that proteins are actively selected and enriched during ER export (active transport). However, the retrieval of HDEL-tagged proteins from the Golgi complex is well established in different cells. The secretion of ER residents is very low, and the true level of ER export is masked by degradation in a post-ER. To overcome this problem, we used two transformed tobacco plants, which produce barley alpha-amylase and alpha-amylase-HDEL as soluble cargo molecules. Labeling using cryo-sectioning and alpha-amylase antibodies revealed that in both transgenic plant cells the secreted molecule was distributed in the ER, cisternae of the Golgi apparatus and cell wall. However, tagging alpha-amylase with HDEL led to predominant labeling of the cis- and cis-most cisternae. Occasional labeling of the compartments distal to the cis-Golgi in alpha-amylase-HDEL–producing plants indicated that saturation of ER retention may have occurred. In addition, both alpha-amylase and alpha-amylase-HDEL were detected in the cell wall, confirming that ER retention via HDEL tagging in the transformed cells was incomplete. Labeling of wild root cells with Calreticulin and Bip antibodies showed that this retrieval mechanism in plants has little impact on ER retention of soluble reticuloplasmins.

Paclobutrazol in one the most important substance from triazol grope. These substance are plant growth retardants which mitigate stress in plants. The aim of the present study was to determine whether the plant growth regulator, such as paclobutrazol can protect tomato seedling from injuries caused by low temperature stress (LTS). In this study two different concentrations (30, 60 mg/l) of PBZ were sprayed on 5-week-old plants. After 3 days, seedlings were exposed to LTS by placing them in cold incubator at 1±0.2 ºC for 6 h for consecutive day and returned to their original place, treatment were applied daily for 5 day. After treatment of seedlings, morphological parameters (such as shoot and root fresh weight, shoot and root dry matter and shoot and root length) and biochemical parameters (such as, proline, leaf chlorophyll, carotenoids, cell membrane lipids peroxidation) were measured. Data of all parameters were analysed with spss software version 9.0. Results showed that LTS inhibited plant growth and induced membrane lipid peroxidation, and increased proline content. On the other hand, LTS decreased the chlorophyll and carotenoids content. PBZ treatment caused shoot dry weight. Internode and shoot length were reduced by this treatment. Instead PBZ, increased root dry weight and root were more extended. Moreover PBZ treated plants when exposed to the cold stress condition, showed reduction in lipid peroxidation and proline content. Treatment of seedlings with PBZ led to an increase in chlorophyll and carotenoid contents either in stress or non stress conditions. Our results showed that PBZ treatment improve chilling injury and responses to the cold stress.

The genus Trifolium L. (Fabaceae) an important member of forage plants comprises approximately 248 species. The germplasm used in this study containing 19 genotypes from 10 species of clover genus considered by Karyotypic studies. In this study, the Video Analysis System was used for karyotype analysis. The basic chromosome number was varied between X=5 (in two diploid genotypes), X=7 (in three diploid genotypes) and X=8 (in twelve diploid genotypes and two tetraploid genotypes). The genotypes number 7585, 27 and 5156 classified to symmetric class of 2A and others stand to 1A. The plot of genotypes, based on two parameters of A1 and A2, and symmetry types of Stebbins had the same results. The Results of analysis of variance based on completely randomized design(CRD) showed a significant differences among the genotypes for all traits (P<%1). Using principal component analysis, the first, second and the third component justify %92.15 of total variance. In the first component, the length of the long arm, long arm relative percentage, short arm, short arm relative percentage and the total length of chromosome which had the highest coefficients of eigen values also had the most significant role in total variance. In the second component the feature of the arm ratio, centromer index and total form percentage had the most important part in creating of total variance. By cutting dendrogram resulted from cluster analysis (UPGMA) based on 9 parameters (TL, LA, %LA, SA, %SA, AR, CI, DRL, %TF) in genetic distance 3.89, the genotypes classified to three classes which certainly the first and the second components had the most significant role in separated classes. In this study there was the highest distance between T.hybridum (27) and T.hirtum(3704) which imply the least affinity between them. There was the least karyotypic difference between two genotypes 314 & 2139 from the species of T.fragiferum. The diagram of the genotypes dispersion, based on two main components, classified the genotypes in three separated groups, supporting the results of cluster analysis. By cutting dendrogram produced from cluster analysis (UPGMA) based on 2 parameters (A1 and A2) in genetic distance 1.75, the genotypes classified to three classes.In this study there was the highest distance between T.hybridum(27) and T.alexandrinum (8313) which imply the least affinity between them. There was the least karyotypic difference between two genotypes T.repens (Alice) and T. subterraneum(7593).

Salicylic acid (SA) is a key endogenous component of local and systemic disease resistance in plants that it is considered as a hormone like substances.A field experiment was conducted during 2004 in research station of Sararod Kermanshah to determine the effects of SA on yield, yield component and resistance of two chickpea cultivars Bivanij (susceptible) and Hashem (resistant) to the pathogen Ascochyta rabiei. The experimental design was a split plot in RCBD with 4 replications in which main plots were 2 cultivars (Bivanij and Hashem) and subplots were 4 concentrations of SA (0, 0.1, 0.7, 1.5 mM).Plant spraying of SA started at beginning of flowering and continued for 20 days. After 10 days of SA treatment, plants were inoculated with Ascochyta rabiei spores suspension (106 spores/ml). The number of primary and secondary branches, plant height, pod length, the number of pods, 100 pod weight, 100 seed weight, and yield per plant were determined. Seed protein was determined by the Bradford method. The results showed that 0.1 mM SA increased yield in both cultivars and increased 100-seed weight and pod length in Bivanij cultivar. 1.5 mM SA significantly decreased total soluble proteins in Hashem cultivar. 1.5 mM SA delayed the onset of disease but at the final stage, disease rating was not decreased significantly.0.7 mM SA had good effects on yield and yield component but increased disease rating in Bivanij. Further investigation on SA effects is needed.

Present study is the faunistics on lizards of Damghan area by analysis of morphological characters and indetification keys. In this research, 231 specimens were collected from different zones of Damghan area during April to October 2004 and transferred to zoological laboratory. Pictures and slides were taken from the live specimens, and then specimens were fixed and preserved in 10% formalin. Identified samples belong to 17 species and 6 families including: Laudakia caucasia, Phrynocephalus maculates, Phrynocephalu scutellatu and Trapelus agilis from the family Agamidae; Ophisaurus apodus from the family Anguidae; Bunopus tuberculatus, Cyrtopodion caspium and Teratoscincus bedriagai from the family Gekkonidae; Eremias fasciata, Eremias intermedia, Eremias persica, Eremias velox velox, Mesalina watsonana and Ophisops elegans from the family Lacertidae; Ablepharus pannonicus and Mabuya aurata transcaucasica from the family Scincidae and Varanus griseus caspius from the family Varanidae. Ophisaurus apodus and Ophisops elegans are reported for the first time in Semnan province. Prior to this research only one specimen of Mabuya aurata transcaucasica had been reported in boundaries between Semnan and Golestan provinces near Shahrood. Here, more specimens are collected from Damghan city.

Effective perception of fear signals is crucial for organism survival. When threated the organism indicated defensive behavior responses that can investigate by using elevated plus–maze. The increas in two parameters of % open arm entries (%OAE) and % time spent in the open arm(%OAT) in the elevated plus – maze were considered as anti –fear effect.In this research the effect of testosterone on fear behavior of gonadectomized rats (GDX), in the elevated plus – maze investigated.. Subcutaneous (SC) injection of different doses of testosterone (100,200,300,450µg /rat) to rats increased % open arm entries (%OAE) and %open arm time spent (%OAT)with dose dependent response manner. Maximum response was obtained with 450µg/rat of the drug (P<0.001).It is concluded that testosterone decreases fear behavior.

Most in vitro studies in the CNS require pure culture of astrocyte.In this article preparation of purified types I and II astrocytes culture from neonatal rat brain cortex were described.The base of this methods is K.D. McCarthy and J.P. Vellis methods (1980)and modified for simplicity of the technique. Separation of the cells from cortex were done mechanical without any enzymes.Single cells were cultured in DMEM+ 10% FCS without enzymes, growth and adhesion factors usage.Astrocyte cultures were characterized as being greater than 99% pure by counting GFAP positive cells over the total number of cells stained by Hoechts #33342. Pure culture of astrocytes can wildly use for different biochemical, molecular genetics, physiology, pharmacology and electrophysiology studies in central nervous system.

The Ghameshloo Wildlife Refuge is situated in 45 km., northwest of Esfahan (in central Iran), and covers a surface of ca. 50,000 hectares. Using physiognomic method, 20 different vegetation units were recognized based on their dominant species. In present work, the vegetation units of the area was examined through establishment of 50 quadrates, within them were recorded the cover of all plant species. In order to assessing the relationships between these quadrates, clustering analysis were applied. The results showed that these quadrates are classified into 2 main clusters and some smaller groups, corresponding to the main vegetation units of the area. The dominant species of these vegetation units are: Scariola orientalis, Astragalus brachyodontus, Astragalus pichleri, Centaurea gaubae, Artemisia sieberi, Artemisia aucheri and Anabasis haussknechtii. Furthermore, the relationships of these vegetation units were discussed.

Diazotrophic cyanobacteria, which are nearly found everywhere, have important role in N fixation and play a vital role in improving and establishing the soils. Little studies have been carried out on the life cycle of heterocystous cyanobacteria in paddy fields of Iran. Sampling, purification and identification of cyanobacteria from paddy fields of Golestan province was carried out and showed that Nostoc ellipsosporum and N.muscorum are the most abundant species of the sites. To draw the growth curves of the species each was grown on Allen culture medium separately and their growth curves were drawn according to their chlorophyll amounts. Doubling time of each species was also calculated based on the chlorophyll data. The results showed that N.ellipsosporum has longer doubling time and logarithmic phase than N.muscorum. Hence, it can have longer activity and fix more N in the ecosystem.