Modares Journal of Biotechnology

Abstract Lipase is used in the production of detergents, cosmetics, pharmaceuticals, flavour enhancers and foods. The lipase of yeast Yarrowia lipolytica can be used for production of important class of chemical intermediates in the pharmaceutical industry. Lipase production depends on media composition and environmental conditions. Y. lipolytica DSM 3286 strain was cultured on media containing different organic and inorganic nitrogen sources. Lipase production was investigated by measuring biomass and lipase activity was detected by ρ-nitrophenyl laurate (PNPL) spectrophotometric assay method at various times within a period of 7 days. In this study, the effect of different nitrogen sources was investigated on Y. lipolytica DSM 3286 lipase production. The maximal lipase production (34.7 U/ml after 48 h) was detected in medium containing yeast extract as nitrogen source. The optimum temperature and pH of the enzyme activity were 37 °C and 7, respectively. The final goal of this study is to develop and optimize lipase production by Y. lipolytica for use in pharmaceutical industry.

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Abstract Lentiviruses are considered one of the most effective recombinant viruses for gene transfer to mammalian cells and tissues. This study comprises of two essential parts: (1) evaluation of efficiency of protein purification columns in concentration of recombinant lentiviruses, and (2) production of recombinant lentiviruses carrying GDNF coding sequences. In part (1) we co-transfected human embryonic kidney cell line HEK-293T with three lentivirus vectors called transfer (carrying either GFP or Jred), packaging and envelope vectors. After a filtration step, we applied the supernatant from transfected cells to Amicon protein columns for concentration purposes. Centrifugation removed 99% of the supernatant and left behind 500-µl-volume of solution full of virions. We thereby produced a of virus stock. Various dilutions of this stock were added to HEK-293T cells that produced up to 100% infected cells positively expressing transgenes. To examine whether the removed supernatant (overflow) has any trace of infective virus by chance, we also used dilutions of the overflow for infection and observed no sign of eGFP or Jred expression. Given the need for a high-titer virus stock for successful target cell transduction, our results indicate that our filtration method of virus concentration is able to produce high virus titer and is cost-effective and less time consuming than previous methods. In part (2), due to the importance of neurotrophic factor GDNF in differentiation and neuroprotection as well as in therapy of neurodegenerative disorders, we ligated GDNF coding sequence into the lentivirus backbone in the second phase of our study. We applied the same method outlined above to produce high-titer recombinant viruses. Following infection of human astrocytoma cells with this virus stock, we detected 3-fold increase in GDNF mRNA expression using RT-PCR. Lentiviruses carrying GDNF can therefore be generated at high titer using the column method and applied for differentiation and neuroprotection studies.

Abstract Abstract: In recent years, extension of Artemia applications in aquaculture and decreasing of natural resource, lead many of related studies to the distribution of Artemia population and new resource assessment studies. Urmia Lake as one the biggest habitat for Artemia because of ecological variations in the regions in which of its differentiations in cyst biometry and Artemia, moreover some genetic variations suggested to have some several Artemia populations in the Lake. In this project Artemia cyst samples were collected from 5 ecological stations of the Lake Urmia. The cyst hatching and the nauplii breeding up to adult Artemia stage were done according to optimum conditions in laboratory. Growth rate and survival of larva in days of 3, 5, 7, 11, 15, 20 of rearing period were measured in feeding with a complex of Dunaliella tertiolecta and Lanzy PZ for a period of 20 days in 75 and 150 ppt. Data and its statistical analysis revealed that according to previously records the growth rate of the Artemia was influenced by increasing the salinity from 75 to 150 ppt survival and growth rate of Artemia have been influenced by water salinity (P<0.05) but increasing the salinity only in two population of Bari and Eslami led decreasing of survival. The produced cysts in two salinity showed that Artemia population can produce the cysts with different diameter and there were not any statistic correlations between the salinity and populations. The Dandrogarm of population statistic analysis emphasized that according to growth rate and survival parameters and among populations under this test there were 4 different populations of Artemia urmiana in which had interesting differentiations in growth rate and survival. Keywords: Artemia, Survival, Growth rate, Salinity, Urmia Lake.

Abstract In order to investigate the Effect of different levels of nitrogen and plant density on yield, and some morphological sunflower cultivars, experimental farm in 2009 in the College of Agriculture University researcher Ardebili A split plot factorial design based on a randomized complete block with three replicates was carried out. Nitrogen treatments included (zero, 75 and 150 kg per ha) assigned to main plots and levels of plant density (8, 10 and 12 plants per square meter) and two varieties were Urofelor and Armavirsky in sub plot. Head and stem diameter, plant height, number of kernel per disk, number of leaves at the final of harvesting and grain yield measured. Sunflower cultivars was significant difference on the stem diameter, plant height and number of kernel per disk but different levels of nitrogen and plant density were significant difference on the for all traits measured. The highest grain yield was obtained, 262 gr /m2 in 150 kg N/ha× Armavirsky. Increasing plant density by increasing the number of heads per unit area, will affect performance. Considering the excellence level of 80 thousand plant density and fertilizer level of 150 kg ha compared to other treatments in terms of grain yield, the amounts of nitrogen and planting density to achieve adequate performance in the test area and similar areas is advisable.

Abstract Atropine and scopolamine are two important tropane alkaloids, widely use as anticholinergic drugs. Such as most solanaceae plants, Atropa belladonna produces these two tropane alkaloids. Carbohydrates are not only act as carbon sources for solanaceus plants in vitro cultures, but also have some effects as signal molecules on tropane alkaloids metabolism. Effect of sorbitol, mannitol and sucrose on growth, chlorophyll, hyoscyamine and scopolamine contents, and hyoscyamine 6- beta hydroxylase (H6H) gene expression, investigated. These treatments decreased the growth of plantlets but increased chlorophyll, hyoscyamine and scopolamine contents. The treatments increase h6h gene expression in different organs of Atropa belladonna plantlets. There was an agreement between h6h expression pattern and scopolamine contents.

Abstract Citric acid is one of the industrial products with extensive applications which are used in the food, pharmaceutical, cosmetics and chemical industries. Although to 1965 Aspergillus niger was single strain to production of citric acid but yeasts are good candidate for citric acid production because growth on cheap and disposal substrates such as hydrocarbon and oils, low sensitive to trace elements in raw material. In this study, citric acid producing yeasts were isolated. Among 340 isolated yeast strains from dairy, meat and food products from Isfahan factories on screening media. 12 strains cultivated in citric acid production medium have been chosen for further study. Production of citric acid was determined by colorimetric method and Megazyme kit during 192 hours. One of the isolated yeasts with 55.5 g/g citric acid production along 144 hours after inoculation had the best yield. Biochemical and molecular tests showed that this strain belonged to the species Yarrowia lipolytica, molecular tests confirmed by sequencing; therefore it was named as Y. lipolytica M7 with accession number HM011048 in Genbank..

Abstract Background: L-lysine is essential amino acid for human and animal nutrition. L-lysine is useful as medicament, chemical agent, food material (food industry) and feed additive (animal food). The industrial production of lysine has become an economically important industrial process. Several hundred thousands tones of L-lysine are produced annually worldwide, almost exclusively using Corynebacterium glutamicum. Study methods: To amplify LysA gene from C.glutamicum, two primers with NheI and HindIII restriction sites were designed. PCR was performed and PCR product was ligated with pTZ57R/T. Recombinant plasmid sequence was determined. LysA with sticky end was ligated with digested pET28a vector and ligation mixture was transformed in E.coli BL21(DE3). The recombinant plasmid was isolated with enzymatic digestion and sequencing. Results: LysA gene, a fragment with 1.3 kb, was cloned. PCR products and enzymatic digestion of extracted vectors with HindIII and NheI, sequencing and SDS-PAGE confirmed the authenticity of cloning. Recombinant bacterial colonies were investigated and confirmed by two methods (PCR and enzymatic digestion). Conclusion: In this study for the first time, the expression rate of Meso- diaminopimelate decarboxylase enzyme (EC 4.1.1.20) in this expression vector was investigated and was increased significantly.