Journal of Sciences, Islamic Republic of Iran
Year:2004 | Volume:15 | Issue:3|Papers Count:18

A self-regulated high-copy number plasmid containing chloramphenicol resistant gene, for the production of recombinant proteins under the regulation of bacteriophage ëpL promoter, was constructed. The designed 5024 base pair expression plasmid contained a heat sensitive repressor cI857 coding gene to regulate the function of ëpL promoter under heat shock induction. Using the constructed vector, for the first time, periplasmic production of human granulocyte macrophage colony stimulating factor (hGM-CSF) under heat induction was demonstrated. The developed hGM-CSF expressing system in this study has potential for the secretion of nearly 100% of expressed proteins into the Escherichia coli periplasmic space upon temperature up-shift.

Chicken infectious bronchitis virus (IBV) causes severe respiratory and renal dysfunction in chickens. In the present study, we used reverse transcriptionpolymerase chain reaction (RT-PCR) to amplify the nucleocapsid (N) protein gene of strain Massachusetts (Mass) H120 of IBV that is commonly used in the vaccine production in Iran. The PCR product with the expected size of 1.2 kb was cloned into a TA-vector and subsequently sub-cloned into expression vector pET- 23a(+). The recombinant plasmids were transformed into competent BL21(DE3) strain of E. Coli. The recombinant N protein was expressed as a soluble fusion protein tagged at its C-terminal with 6 x His residues. In Western blot analysis, a convalescent serum from a chicken tested positive by a commercial IBV detection kit specifically reacted with the recombinant protein. The IBV negative chicken serum did not react with the recombinant protein. In a reciprocal experiment, antiserum raised against the purified fusion protein reacted with the corresponding proteins of strains H120 and 4/91. This is the first report of cloning and expression of the N protein gene cDNA of strain Mass H120 of IBV in a bacterial expression system. The potential uses of the cloned gene and the recombinant N protein have been discussed.

Leukemia is referred to a group of hetrogenous neoplasia, which results from the hematopoietic cells transformation. Various chromosomal abnormalities have been recognized as primary patogenetic changes. Clonal chromosomal aberration is found in approximately 80% of childhood acute leukemia. Favourable and poor prognosis karyotypes are evident in such patients. Since cytogenetic play a dominant role in outcome of patients with childhood acute leukemia it is important to associate characteristic cytogenetic with prognostic value in relation to chromosomal abnormalities. Cytogenetic analysis from 51 childhood diagnosed patients under the age of 14 has been performed on the basis of G-banding technique. Each pair of chromosome show unique banding pattern. Various chromosomal abnormalities were evident in this study, these include; numerical disorders such as: monosomy, trisomy, tetrasomy, pentasomy and pseudodiploidy. Among them monosomies were more prominent than the other aneuploidy. Structural aberrations such as philadelphia chromosome, gain and loss, inversion and breakage were seen. Each kind of childhood acute leukemia shows different responses to chemotherapy following cytogenetic studies. This finding strongly suggests that, a close association exists between chromosomal abnormalities and the prognostic value and prediction of outcome.

Helicobacter pylori is the causative agent of chronic gastritis and peptic ulcer diseases and is also a risk factor for gastric cancer. Since culture of Helicobacter pylori is relatively insensitive and cumbersome, PCR-based molecular detection and typing of this organism is gaining importance for strain differentiation. The aim of present study was to apply molecular methods for detection and genotyping of H. pylori directly on gastrointestinal biopsy. For this purpose 79 samples were tested by bacteriological methods including Gram stain, urease test and culture in a selective medium. Genomic DNA was directly extracted from gastric biopsy specimens followed by PCR amplification using ureC primers. PCR products from positive samples were purified and digested using three different restriction endonucleases, AluI, MboI and CfoI. The numbers of positive samples detected by bacteriological methods were 71, 40 and 73 using Gram stain, culture and urease test respectively. Through PCR assay 64 samples were positive with sensitivity of about 92 and specificity of 100%. The specificity and sensitivity of the PCR assay was higher than that of all bacteriological methods applied. RFLP performed on the positive samples and five patterns for AluI, five patterns for MboI and four patterns for CfoI detected. The patterns were classified based on the number of produced bands in gel electrophoresis. From our study it is concluded that the PCR-based RFLP technique would be an useful approach for identification and genotyping of specific H. pylori strains directly in gastric biopsy specimens without culture.

The PML gene encodes a nuclear protein with transcriptional regulatory activity. It directly interacts with the Sp1 transcription factor and abrogates its activation of the epidermal growth factor receptor (EGFR) gene promoter. Here, the effects of PML on the activity of dihydrofolate reductase (DHFR) promoter, mainly regulated by Sp1, were investigated. On functional analysis, transient transfection of PML into mammalian cells, U2Os and HeLa, resulted in a significant repression of the DHFR promoter. The DHFR promoter also contains binding sites for the E2F transcription factor. When the Sp1 DNA-binding site, but not the E2F-binding site of the promoter, was replaced with an unrelated binding site (GAL4), the repressive effects of PML were lost. Moreover, electrophoretic mobility shift assay (EMSA) using Sp1-containing oligonucleotide probes showed significant reduction in Sp1 binding in the presence of PML, confirming the necessity of the Sp1 DNA-binding sites for PML’s repressive effects. Analysis of DNA synthesis using [3H]thymidine incorporation assay showed a significant reduction in DNA synthesis in HeLa cells overexpressing PML. Together, the data demonstrated that PML could function as a negative regulator of the DHFR promoter, which may represent a novel mechanism for the known repressive effects of PML on cellular growth.

Among the different reducing agents tributyltin hydride was found to be a powerful reducing agent for the denitration of a series of tertiary and secondary nitroesters. The organotin residues removed from the crude product using saturated potassium fluoride solution. The yield of the reactions was 62-98%.

The molecular associations in the binary mixtures of (methanol/solvent), (ethanol/solvent) and (propanol/solvent) were studied using NMR, FTIR and VLE data at different temperatures. The activity coefficients as the most important quantities representing the mixture deviation form ideal behavior were evaluated by using various association models to express the physical and chemical contributions of the activity coefficients. The parameters of these models were calculated. The results indicated that for methanol and ethanol the dominant associated species are the tetramer whereas for propanol it is the trimer. The differences between the parameters of activity coefficient models were calculated by considering the physical and chemical contributions separately and were expressed in terms of residual parameters for the activity coefficient models. The parameters of van Laar model were corrected on the basis of the existing dominant species in the associated mixtures and a simple linear relation was obtained for the parameters of the model for the dominant species in terms of the parameters for monomer.

The degree of carbonate cementation in argillaceous sediments represents one of the significant diagenetic evolutions in ancient mudstones. The alternation of the black shale and laminated limestone of the Lower Jurassic of SW Britain is an example, which typify extent of diagenetic modification of marine clay and organic-rich sediments. The Lower Jurassic mudstones of the SW Britain include black shales, marls and alternate with limestone beds and nodules. Laminated limestones are also present and are only enclosed between the black shales. Detailed petrography combined with geochemical analysis of the black shales and enclosed laminated limestoned have been carried out and compared to the typical isolated nodules in order to interpret the diagenetic history of mudstones. Displacive growth of the microspar crystals was an important process that occurred during the diagenetic evolution of these mudstones. The black shales show carbon (+0.9 to +1.9‰ (PDB)) and oxygen (-2.2 to -4.1‰ (PDB)) isotopic values, which is compared to their enclosed laminated limestones, that show more negative carbon (-0.2 to -1.2 ‰ (PDB)) and oxygen (-5.4 to -6.8‰ (PDB)) values. The stratigraphic position, petrography and isotopic relationship of the black shales and enclosed laminated limestones indicate a degree of diagenetic evolution and carbonate cementation of mustones during progressive burial.

The empirical Bayes estimators of treatment effects in a factorial experiment were derived and their asymptotic properties were explored. It was shown that they were asymptotically optimal and the estimator of the scale parameter had a limiting gamma distribution while the estimators of the factor effects had a limiting multivariate normal distribution. A Bootstrap analysis was performed to illustrate the theoretical results empirically.

Copper films of 300 nm thickness deposited by planar magnetron sputtering on  glass substrates, for substrate temperatures between 300 and 500 K, and deposition rate of 10 Ås−1. Microstructure of these films was obtained by X-ray diffraction, while the texture mode of diffractometer was used for stress measurement by the sin2ψ technique. The components of the stress tensor are obtained using measurements at three different φ angles of 0, 45 and 90 degrees by proposed technique in this work. The relation between stress in these films and the processes in film growth through structure zone model (SZM) is explained.