Showing 3 results for Rahmani
Mohammad Hassan Kheirandish, Hossein Zarei Jaliani, Behnaz Rahmani,
Volume 4, Issue 3 (August 2017)
Abstract
Background and Aims: DNA cloning, sub-cloning and site directed mutagenesis are the most common strategies in nearly all projects of recombinant protein production. The classical method of restriction site cloning is unsatisfactory due to the need for supply of restriction enzymes and the inefficiency of the digestion reaction. Many new methods, including recombinatorial cloning and ligation independent cloning need additional enzymes and kits. In this project we insert a full-length streptolysin O gene into an expression plasmid without using any uncommon commercial enzymes.
Materials and Methods: Steptolysin O gene was amplified by polymerase chain reaction (PCR) and introduced into the pPSG-IBA35 vector using a quick-change PCR. At the same time the gene was double digested and sub-cloned into pET28a (+). Both constructs were introduced into BL21 DE3 cell. Proteins were purified by Ni-NTA column and hemolytic activity was evaluated by spectrophotometry using human red blood cells.
Results: Steptolysin O was subcloned into the pET28a (+) and pPSG-IBA35 vectors and expressed in E. coli. Protein was purified with over 90% purity. The IC50 of C and N terminus his-tagged protein were 0.22 and 0.29 µg/ml, respectively in hemolysis assay.
Conclusions: This study showed for the first time that full-length streptolysin O can be expressed in E. coli cytoplasm without any toxicity for the bacteria itself. The only additional amino acids expressed on the protein were his-tag. To study the role of this toxin it would be better to express the protein with the same strategy to have minimal extra amino acids on the protein.
Behnaz Rahmani, Hossein Zarei Jaliani, Akram Astani, Mohammad Hassan Kheirandish, Ahmad Mosaddegh, Abolghasem Asadi Saghandi,
Volume 4, Issue 4 (November 2017)
Abstract
Background and Aims: Enzybiotics are probably the future line of weapons against drug resistant bacteria. They lyse bacteria with the new mechanisms with few likelihood of generating resistance. LasA, which is secreted from Pseudomonas strains degrades Staphylococcus aureus (S. aureus) cell wall and has the potential to use against drug resistant S. aureus infections.
Materials and Methods: Codon-optimized gene of the mature form of the LasA protein was ordered. The gene was double digested with NcoI and XhoI restriction enzymes and sub-cloned into pET28a(+) digested with the same enzymes. Recombinant construct was introduced into BL21 DE3 cell. Expression of the gene was induced by 0.2 mM isopropyl β-D-1-thiogalactopyranoside and recombinant protein was affinity purified by Ni-NTA mini-column. The staphylolytic activity of the recombinant LasA protein was evaluated on Methicillin-resistant S. aureus (MRSA) by disk diffusion.
Results: Fragment of the LasA gene encoding mature form of the LasA protein was introduced into pET28a(+) expression vector. C-terminal
his-tagged recombinant LasA protein was produced in BL21 DE3 E. coli cells. Over 50% purity has been achieved by affinity purification of the LasA protein from the total cell lysate. The yield of purified protein was 5.4 mg.l-1. Growth of MRSA was completely inhibited by dilutions of recombinant his-tagged LasA.
Conclusions: The present study shows that the mature form of the LasA can be expressed in E. coli BL21 DE3 cells. C-terminal his-tagged form of the mature LasA protein has staphylolytic activity against MRSA and so it can be a promising therapeutic agent.
Mostafa Amiri, Mohammadreza Rahmani, Hossein Solemani, Mohammad Ghorbani, Roghaieh Rahmani Bilandi,
Volume 5, Issue 2 (May 2018)
Abstract
Background and Aims: Preeclampsia is one of the causes of maternal mortality whose leading cause is still unknown. This study was conducted to investigate the relation between blood group and pre-eclampsia in primipara women.
Materials and Methods: This case-control study was performed on 100 pirmipara women in Gonabad city. The samples were assigned, by census, into two groups of case (with pre-eclampsia) and control (without pre-eclampsia). The data were analyzed using descriptive statistics and Chi-square through SPSS software Ver.16.
Results and Conclusions: The incidence of preeclampsia in the blood group of A, B, AB, and O were 36%, 34%, 22%, and 8%, respectively. Also, there was a significant difference between blood group O and blood group non-O to preeclampsia (p<0.001). The findings showed that the women with A and B blood types require special care with an emphasis on controlling and preventing the factors which cause hypertension/preeclampsia during pregnancy.
