Journal of Molecular and Cellular Researches
Year:2020 | Volume:33 | Issue:1|Papers Count:12

In order to increase the availability of phosphorus and other minerals in the diet of birds and monogastrics and reduce environmental pollution this reasearch was aimed to produce phytase. This was designed to produce a suitable enzyme for digestion of phytate in the gastrointestinal tract of birds. For this propose, Aspergillus niger phytase was produced on PSM media and Escherichia coli and Bacillus subtilis phytases with using recombinant DNA technology. The enzymes were purified by Q-Sepharose and Ni-NTA chromatography column and, using SDS-PAGE, their molecular weights were estimated 42 kDa, 44 kDa and 66 kDa for PhyC (Alkaline Phytase), appA (Escherichia coli Phytase) and PhyA (Acid Phytase) respectively. The phytase activity for PhyC, appA and PhyA were determined 54. 81, 35. 49 and 29. 33 U/ml, respectively. Comapring their kinetics parameters, it was shown that PhyC enzyme can be active in 90 0 C and had thermostability while appA enzyme had best rang of activity in pH of 4-4. 5 and efficient enzyme in gastrointestinal tract and appA had maximum affinity to substrate by km 0. 09± 0. 03. According to the results, a suitable combination of these enzymes could be efficient enough to hydrolyse phytate in gastrointestinal tract of birds.

Malva is one of the well-known medicinal and aromatic plants. The juice is one of the oldest herbs that are widely used in medicine. DNA extraction was carried out using CTAB method for 19 genotypes of different regions of Iran. Using SCoT molecular markers, genetic diversity was investigated. The SCoT marker showed favorable polymorphism among genotypes and most primers were suitable for this species. SCoT primers were able to produce 83 strips in total, with 76 multidimensional bands. The average number of bars produced by each primer for 19 genotypes was 4. 36, with SC36, SC11 and SC5 primers having the highest number of bands (8), and the SC26 and SC15 primers showed the least number of bands (3). The results showed that the most similarities were genotypes G18 and G19 and the least similarity was genotypes G15 with G3 genotype. The cluster analysis showed that the genotypes were classified into three groups and genetic variation was not consistent with geographical variation. The results of cluster analysis were grouped using molecular variance analysis (AMOVA).

L-DOPA (L-3, 4-dihydroxyphenylalanine) was first discovered in the 1960s as the most effective treatment for Parkinson's disease. In the present research, for the first time, isolation and identification of L-tyrosine-degrading moderately halophilic bacteria and evaluation their ability as biocatalyst for the bioconversion of L-tyrosine into L-dopa was studied. Using an enrichment technique in tyrosine-containing medium, the Ltyrosine-degrading strain SL-7 with the highest ability to bioconvert L-tyrosine into Ldopa was phenotypically and molecularly characterized and its 16S rDNA sequence was submitted as Halobacillus sp. SL-7. In order to increase the reaction efficiency and prevent the degradation of the metabolites (L-dopa), One-factor-at-a-time optimization method was used to improve the yield of L-dopa in the conversion reaction under resting cells of Halobacillus sp. SL-7. Based on our results, the optimum bioconversion conditions for the production of L-dopa can be summarized as follows: initial tyrosine concentration 1. 5 g/l, biomass concentration 7. 5 g/l, copper concentration 0. 03 g/l and peptone at the concentration 1 g/l as cosubstrate. Under these conditions, the maximal L-dopa concentration (0. 75 g/l with a molar yield of 46. 2%) was achieved after a 36 h reaction. This is the first report on the potential of resting cells of Halobacillus sp. SL-7, A moderately halophilic bacterium with ability biological converting L-tyrosine into Ldopa.

Fusarium seedling blight is a disease of cereal crops caused by a group of trichothecene producing Fusarium species such as Fusarium graminearum. Control of seedling blight has received less attention than control of Fusarium head blight. Application of chemicals which activate plant defense mechanisms before pathogen attack without environmental side effects of protective chemical agents have stimulated great deal of researches in this area. Hence in this study, the effect of seed priming with salicylic acid on Fusarium seedling blight incidence and severity was investigated. According to our results seed priming with salicylic acid improved wheat defense against Fusarium seedling blight in both cultivars. Salicylic acid treatment induced chitiniase gene expression in both cultivars as compared to their controls; however, this increase was much higher in salicylic acid treated Falat than salicylic acid treated Sumai3. Based on the results, increase in gene and protein expression of chitinase could induce host resistance against Fusarium seedling blight caused by F. graminearum.

Bread wheat is the most important crop in the world whose yield is affected by several diseases including bacterial leaf blight. The most appropriate method for managing this disease is using resistant cultivars. DNA markers are highly effective in plant grouping in terms of resistance to diseases, and among these markers, the ISSR marker is recommended because of the do not need for prior information from the DNA sequencing and its easy and fast implementation. In this research, genetic variation of 27 Iranian indigenous wheat cultivars with respect to resistance to bacterial blight was investigated using 12 ISSR primers, which produced 170 evaluable bands, of which 156 were polymorphic bands. The highest percentage of polymorphic bands was belonged to F7, F8 and F10 primers, and the lowest of it was related to F9 primer. The highest content of the polymorphism information content (PIC) was belonged to the F10 and F5 primers and the smallest value belonged to the F7 primer. Cluster analysis classified these cultivars into four groups: Ghods, Omid and Atrak cultivars in the cluster of resistant group and other cultivars in sensitive and very sensitive group. The high polymorphism obtained in this study can be attributed to the high efficiency of the ISSR marker in evaluating the genetic diversity of wheat cultivars in terms of resistance to various diseases.

β-Glucans (beta-glucans) form a natural component of the cell walls of bacteria, fungi, yeast, and cereals such as oat and barley. Glucanases are enzymes that break down a glucan. β-glucans are chains of D-glucose polysaccharides linked by β-type glycosidic bonds. The purpose of this study is to express the lechinase gene in bacteria to produce recombinant enzyme as a feed supplement in poultry diets. In this study, LicBM2 gene, isolated from Clostridium thermocellum, encodes thermostable lichenase enzyme was cloned in expression vector pET22b (+) and E. Coli bacteria strain BL21. Bacterial β 1, 3– 1, 4-glucanases (EC 3. 2. 1. 73; lichenase) specifically cleave β-1, 4-glycosidic linkage adjacent to 3-O-substituted glucopyranose residues. Gene expression was confirmed using SDS-PAGE techniques. The enzyme activity of recombinant protein and reducing glucan in barly were measured by DNS method. The optimum temperature for recombinant protein production was 55 ° C and the maximum enzyme activity was obtain within 4 hours after inducing in pH = 8. Using barlay in poultry diet is more economical. But the significant amounts of beta-glucans in barlay cause more problems. The recombinant enzyme produced in this study can be used as a feed supplement for hydrolyzation of barley β-glucan to replace of corn by barley in poultry diets.

Genetic manipulation of crop plants to obtain resistance to herbicide glyphosate is one of the most effective approaches for weed management. Application of the genes encoding glyphosate degrading enzymes such as glyphosate oxidoreductase (GOX) in combination with a glyphosate-tolerant EPSPS is the ultimate approach to provide commercial rates of glyphosate tolerance. The gox gene was first isolated from Ochrobactrum antrophi strain LBAA that catalyses the cleavage of glyphosate into aminomethylphosphonic acid and glyoxalate. In this study, the gox gene was synthesized and cloned in pET28a expression vector and transformed in E. coli. The expression of the target protein was confirmed by SDS-PAGE. The experiment for optimization of gene expression showed that the optimal condition was 37 o C for bacterial culture, 4 hours of induction time using 1 mM IPTG in OD600=0. 6. In the next step, in order to perform a bioassay on transformed bacteria, the threshold of tolerance of control bacteria in minimum phosphate-free medium with different concentrations of glyphosate was investigated and compared with recombinant bacteria. The result indicated that the wild type bacteria were not able to tolerate concentration of more than 0. 5 mM glyphosate but the recombinant bacteria survived to a concentration of 1 mM. These results were confirmed by colony counting on selective media with three repetitions. Hence, glyphosate oxidoreductase gene could be a suitable candidate besides the other glyphosate resistance genes for genetic manipulation of strategic plants with the aim of obtaining higher and more sustainable levels of resistance to this herbicide.

Medicinal plants of the Asteraceae family are important genetic resources due to their high echological flexibility in diverse climates, . Yarrow (Achillea millefolium subsp. millefolium) is a self-pollinated plant belongs to the Asteraceae which produce a wide varieties of plant secondary metabolites such as terpenoids and phenylpropanoids. In the present work, gene expression patterns of genes involved in the biosynthetic pathways of terpens and phenylpropanoids were studied to understand the regulatory mechanism behind them. 1-deoxy-D-xylose-5-phosphate reductoisomerase (DXR) and geranyldiphosphate synthase (GPPS) are important genes in the biosynthesis of monoterpenes in the 2-C-methylerythritol-4-phosphate (MEP) pathway. Phenylalanine ammonia lyase (PAL) and chalcone synthase (CHS) genes are important in the biosynthesis of phenylpropanoids. In the present study, semi quantitative RT-PCR of these genes was carried out in different developmental stages, also in response to methyl jasmonate treatment in leaves and flowers. Results showed that expression of these genes were higher in flowers and methyl jasmonate treated plants, also the expression of these genes affected by developmental stages of leaves but with different intensity.

Cucurbita genus belongs to Cucurbitaceae family. These plants are attacked with different pathogenic agents. Currently the effective strategy to control many disease of Cucurbitaceae is the usage of resistant plants. In this study, the NBS domain encoded by resistance genes was studied in Iranian native varieties of cantaloupe. For this purpose, the seeds of Iranian native cantaloupe cultivars were provided and cultivated in greenhouse. DNA was extracted from leaves of different cultivars using CTAB method. Degenerate primers were designed from conserved motives of resistance analogues genes and amplification of these genes was performed using PCR method. The results showed the isolation of an analogous for resistance gene to tomato mosaic virus (ToMV) in cantaloupe cultivars TN-92-99 and TN-92-80 that were cloned in pGEM-T plasmid and then sequenced. Blast analysis indicated that NBS sequence in Irnian cantaloupe share 92% similarity with ToMV resistance gene in watermelon. Phylogenetic tree drawing using resistance gene analogues from cantaloupe and watermelon existing in NCBI gene bank created tow main groups and seven subgroups. Second structure study of protein of ToMV resistance gene analogus using PSIpred software indicated that these proteins only have α helix structure. This protein is secreted in plasma membrane of cantaloupe cultivar TN-92-99 and its extracellular secretion is very little. This is the first study of cloning, expression and identification of enzymatic activity of a resistance gene against ToMV in Iranian native cantaloupe cultivars. Expression of this gene could play important role in resistance against viral diseases including tomato mosaic disease in Cucurbitaceae that could use in breeding programs.

Iran ranks the third one in the world for the production of almond and over 30 Amygdalus (Prunus) species, subspecies or ecotypes have been identified in the country. Among the wild almond species, P. scoparia is used as the rootstock for almond in southwest of Iran. This species provides a gene pool of valuable characteristics for breeding and improvement of the cultivated almond. The present study considers genetic diversity analysis of three P. scoparia populations by using ISSR markers. We tried to show genetic divergence versus admixture of these populations. ISSR loci showed 50% polymorphism among 38 wild almond trees studied. The highest values for the genetic diversity, shannon index and the percentage of polymorphism were observed in Deihook (Yazd) population. In cluster analysis, Deihook population stands much separated from the other two populations while, Rashm-Moalem (Semnan) and Fasa (Fars) populations showed a higher genetic exchange among their trees. AMOVA test showed great genetic differences among the studied populations. The Mantel test did not show an association between the genetic and geographic distances of the studied populations. This is possibly due to gene exchange among these populations which was also shown by reticulation and STRUCTURE analyses.

Polycyclic aromatic hydrocarbons (PAHs) are an important group of pollutants which are released into the environment mainly due to incomplete combustion of organic materials. PAHs frequently detected at relatively high concentrations in soils of some regions and are toxic for organisms including plants. In this study, wheat plants were grown hydroponically and treated by different concentrations of phenanthrene and pyrene. After determination of effective levels of two compounds, their effects on some physiological and biochemical parameters were evaluated. While pyrene showed no significant effect on studied parameters, phenanthrene significantly decreased photosynthetic pigments content and membrane stability index of shoot and root. Also, phenanthrene significantly increased malondialdehyde content of shoot and root; and hydrogen peroxidase content of root but decreased hydrogen peroxidase of shoot. Phenanthrene significantly decreased CAT, POD, SOD and APX activity in shoot, while pyrene only decreased APX activity. Both compounds decreased CAT, POD, SOD and APX activity in root, but the effect of phenanthrene was higher than that of pyrene. According to the results of this study, phenanthrene inhibited antioxidant enzymes activities and caused oxidative stress more than that of pyrene, and therefore led to higher reduction in photosynthetic pigments, plasma membrane integrity and ultimately plant growth.

Drought stress causes oxidative damages to the macromolecules, such as proteins, lipids and nucleic acids. Two of compounds which has antioxidative characteristic is salicylic acid (SA) and ascorbic acid (As) can decrease drought effects in plants under stress. In recent research, we evaluated parameters such as; changes in activity of catalase and peroxidase and flavonoids in the Hibiscus esculentus L. under drought stress conditions. The results was showed that flavonoids and peroxidase activity increased compared to control under drought stress conditions. Flavonoids and peroxidase activity decrease in plants were under salicylic acid, ascorbic acid and both of them treatment. Catalase activity in drought stress conditions decrease. Catalase activity increased in plants were under Salicylic acid and ascorbic acid treatment. Based on our results, it seems that salicylic acid and ascorbic acid can regulate the amount of flavonoids and the activity of antioxidant enzymes such as catalase and peroxidase in order to reduce the oxidative stress caused by drought. Therefore, the use of these compounds in drought conditions is recommended.