hossein aghamollaei

Sulfur mustard (SM) is a potent blistering agent. This alkylating chemical agent has extremely toxic effects on the eye. MMP-2 and MMP-9 are the two most important matrix metalloproteinase enzymes involved in the pathology of chemical eye injuries. Curcumin is regarded as a natural anti-inflammatory agent. This study aims to compare the anti-inflammatory effects of curcumin versus doxycycline on chemically induced corneal injuries.

The HCE-2 cell line was used as a model for corneal cells. The effective concentrations of 2-chloroethyl ethyl sulfide (CEES) – as an analog of SM – doxycycline, and curcumin were determined using the MTT assay. The gene expression of MMP-2, MMP-9, and tissue inhibitors of metalloproteinase (TIMP-1) was evaluated by the real-time PCR method. Also, the activity of MMP-2 and MMP-9 enzymes was determined by zymography.

The expression of the MMP-2 and MMP-9 genes increased 5- and 3.3-fold after exposure to CEES, respectively. Following the treatment with curcumin and doxycycline, MMP- 2 expression decreased significantly. Also, after treatment with curcumin and doxycycline, the MMP-9 expression decreased 2.5- and 1.6-fold, respectively. The reduction in activity was 32% for MMP-2 and 56% for MMP-9 after treatment with curcumin. The corresponding values were 12% and 40% following doxycycline treatment. There was no significant difference between the effects of curcumin and doxycycline on reducing MMP-2 expression, but the difference was statistically significant in the case of MMP-9.

Doxycycline and curcumin can inhibit MMP expression and activity in chemically exposed corneal cells. Curcumin has a greater ability than doxycycline to inhibit MMP-2 and MMP- 9 enzymes; however, the difference is statistically significant only in the case of MMP-9. After further validation, these substances can be introduced as anti- inflammatory agents to treat corneal chemical burns.

The epidemiology of ocular manifestations and associated complications in patients with Corona Virus Disease 2019 (COVID-19) have been investigated worldwide. Herein, we aimed to summarize the frequency of ocular symptoms of COVID-19 in the current systematic review and meta-analysis using available literature.

A search of PubMed, Scopus, Web of Science, EBSCO, and Embase electronic databases to review the systematic literature until August 2021. The Hoy et al., 2012 tool was used to evaluate the quality of studies. For Data extraction, two reviewers blind and independently extracted data from the abstract and full text of the studies included. 95% confidence interval for effect size with random effect model and restricted maximum likelihood (REML) method were calculated. Meta-analysis was performed using Stata/MP v.16 software (The fastest version of Stata).

Initially, 210 studies were retrieved, of which we reviewed the full text of 24 records, and eight were selected for final meta-analysis. The prevalence of redness in patients with COVID-19 was 12.37% (ES, 95% CI -1.11%, 25.85%), and heterogeneity was found (I2 = 99.37%; p <0.001) (High heterogeneity). The prevalence of Ocular manifestations as the first symptom of COVID-19 in patients was 2.63% (ES, 95% CI 1.23%, 4.03%).

This systematic review and meta-analysis found the prevalence of redness, dryness, ocular pain, foreign body sensation, discharge, itching, follicular conjunctivitis, and watering as 12.37%, 26.70%, 30.64%, 14.47%, 19.93%, 8.77%, 12.77%, and 10.23%, respectively.

Treatment of malignancies with chemotherapy and surgery is often associated with disease recurrence and metastasis. Immunotherapy improves cancer treatment by creating an active response against tumor antigens. Various cancer cells express a large amount of glucose-regulated protein 78 (GRP78) protein on their surface. Stimulating the immune system against this antigen can expose cancer cells to the immune system. Herein, we investigated the effectiveness of a cGRP78-based vaccine against different cancer cells.

Experimental approach: 

BALB/c mice were immunized with the cGRP78. The humoral immune response against different cancer cells was assessed by Cell-ELISA. The cellular immunity response was determined by splenocyte proliferation assay with different cancer antigens. The effect of vaccination on metastasis was investigated in vaccinated mice by injecting melanoma cancer cells into the tail of mice.

These results indicated that the cGRP78 has acceptable antigenicity and stimulates the immune system to produce antibodies. After three injections, the amount of produced antibody was significantly different from the control group. Compared to the other three cell types, Hela and HepG2 showed the highest reaction to the serum of vaccinated mice. Cellular immunity against the B16F10 cell line had the best results compared to other cells. The metastasis results showed that after 30 days, the growth of B16F10 melanoma cancer cells was not noticeable in the lung tissue of vaccinated mice.

Conclusion and implications: 

Considering the resistance of vaccinated mice to metastasis, this vaccine offers a promising prospect for cancer treatment by inhibiting the spread of cancer cells.

Breast cancer is one of the most prevalent malignancies in women. Several treatment options are available today, including surgery, chemotherapy, and radiotherapy. Immunotherapy, as a highly specific therapy, involves adaptive immune responses and immunological memory. In our present research, we used the recombinant C-terminal domain of the GRP78 (glucose- regulated protein 78) protein to induce an immune response and investigate its therapeutic impact in the 4T1 breast cancer model.

BALB/c mice were immunized with the cGRP78 protein. The humoral immune response was assessed by ELISA. Then, BALB/c mice were injected subcutaneously with 1×106 4T1 tumor cells. Subsequently, tumor size and survival rate measurements, MTT, and cytokine assays were performed.

The animals receiving the cGRP78 vaccine showed significantly more favorable survival and slower tumor growth rates compared with unvaccinated tumor-bearing mice as the negative control mice. Circulating levels of tumoricidal cytokines such as IFNγ were higher, whereas tolerogenic cytokines such as IL-2, 6, and 10 either did not increase or had a decreasing trend in mice receiving cGRP78.

cGRP78 vaccines generated potent immunotherapeutic effects in a breast cancer mouse model. This novel strategy of targeting the GRP78 protein can promote the development of cancer vaccines and immunotherapies for breast cancer malignancies.

Recent studies imply extensive applications for the human amniotic membrane (hAM) and its extract in medicine and ophthalmology. The content of hAM meets many requirements in eye surgeries, such as refractive surgery as the most important and commonly used method for treating the dramatically increasing refractive errors. However, they are associated with complications such as corneal haziness and corneal ulcer. This study was designed to evaluate the impact of amniotic membrane extracted eye drop (AMEED) on Trans-PRK surgery complications.

This interventional and historical study was performed during two years (July 1, 2019-September 1, 2020). Thirty-two patients (64 eyes), including 17 females and 15 males, aged 20 to 50 years (mean age of 29.59 ± 6.51) with spherical equivalent between -5 to -1.5 underwent Trans Epithelial Photorefractive Keratectomy (Trans-PRK) surgery. One eye was selected per case (case group) and the other eye was considered as control. Randomization was done using the random allocation rule. The case group was treated with AMEED, and the artificial tear drop every 4 hours. The control eyes received artificial tear drops instilled every 4 hours. The evaluation continued for three days after the Trans-PRK surgery.

A significant decrease in CED size was found in the AMEED group on the second day after surgery (P=0.046). Also, this group had a substantial reduction in pain, hyperemia, and haziness.

This study showed that AMEED drop can increase the healing rate of corneal epithelial lesions after Trans- PRK surgery and reduce the early and late complications of Trans-PRK surgery. Researchers and Ophthalmologists should consider AMEED as a selection in patients with persistent corneal epithelial defects and patients who have difficulty in corneal epithelial healing. We understood AMEED has a different effect on the cornea after surgery; therefore, the researcher must know AMEED’s exact ingredients and help expand AMEED uses.

Keratocytes are the major components of the human corneal stromal cell. Cell therapy by keratocytes can be used in some corneal diseases. Because keratocytes are mitotically quiescent; therefore, the cultivation of these cells is associated with challenges. The present study aimed to isolate, culture, and validate keratocyte cells from discarded corneal tissue based on optimizing some cultivation conditions. In this experimental study, keratocytes were isolated from discarded corneal tissue. Different culture medium composition such as amniotic membrane extract, time, and the role of coating scaffolds was evaluated. Real-time PCR of specific genes were used to confirm the primary keratocyte cells compared to corneal epithelial cells. The specific genes were keratocan, lumicane, aldehyde dehydrogenase three members of family A1 (ALDH3A1), and CD34. In addition, immunocytochemistry (ICC) was used to confirm the expression of specific keratocan and lumican markers. Keratocytes was isolated and cultured in the culture medium containing amniotic membrane extract. Based on analyses, keratocan, lumicane, ALDH3A1, and CD34 gene expression in keratocytes was significantly higher than in the epithelial cells. Moreover, keratocan and lumican expression was detected in 92.5% and 91.1% of the cells, respectively. According to the results, the addition of amniotic membrane extract significantly increased the growth of keratocytes. Our findings in this study showed that discarded corneal tissue can be used as a suitable source for obtaining keratocyte cells needed in corneal tissue engineering.

Keratopathy is a common complication following exposure to chemical warfare, namely mustard gas. In recent years, a quest was undertaken to seek an effective and uncomplicated corneal transplantation method by ophthalmologists nationwide. In our study, we report our results following lamellar keratoplasty (LK) without limbal stem cell transplantation in patients with delayed-onset mustard gas keratopathy (DMGK).  Thirty-one eyes of 22 veterans with DMGK who underwent conventional LK were assessed. The presence of limbal stem cell deficiency (LSCD) was proven clinically as well as by impression cytology. The results were evaluated based on best spectacle-corrected visual acuity (BSCVA), refractive error (RE), corneal clarity, and corneal graft survival rate. The mean age of patients at the surgery was 52.93±5.8 years, and the mean follow-up was 36.25±20.65 months. The mean preoperative BSCVA was 1.67±0.32 logarithm of the minimal angle of resolution unit (Log MAR), significantly improved, reaching 0.32±0.22 Log MAR after surgery (p = 0.001). The mean preoperative spherical equivalent (SE) was 2.47±1.17 diopters, significantly increasing to 3.5±1.08 diopters after suture removal was complete (p = 0.001). Epithelial graft rejection occurred in 9.67% of eyes (3/31) and was treated successfully with topical eye drops. At the end of the follow-up period, no signs of graft rejection were found, but three eyes had mild central corneal opacity. According to the results of our study, LK without LSC transplantation can be successful in patients with DMGK because of partial rather than complete LSCD.

Delayed mustard gas keratopathy (DMGK) is the main chronic outcome in eye-chemical injured patients. The aim of this study was the histopathological evaluation of mustard-exposed cornea after more than 30 years.

Fourteen corneas after Lamellar keratoplasty were evaluated in this study. Corneal tissues were prepared by histologic methods and stained by H&E.

The main histopathological findings in these cases were the presence of severe stromal edema and corneal scar. In the sections with visible superficial epithelium, subepithelial bullae formation was observed. Focal or diffuse disruption of Bowman's membrane and replacement with fibrosis were also seen. There was no evidence of stromal vascularization and inflammation in all specimens.

After more than 30 years, an extensive corneal scar is seen in sulfur mustard exposed patients. Scar tissue without vascularization and fibroblastic proliferation is the main finding in the sulfur mustard exposed cornea. This pathology result is different from other scars. No evidence of inflammation or immune cell infiltration should be considered in managing DMGK.

Eye injuries are one of the most common side effects of chemical agents, especially mustard gas, which can cause many conflicts in the long time. The cornea is the most critical component of the eye, and many eye injuries somehow are related to that. Nowadays, corneal transplantation is the conventional treatment for corneal injuries, which is due to the lack of a donor, rejection of the transplant, and the possibility of transmitting the disease from the donor to the recipient, face limitations that require an alternative with high efficiency. Stem cells with the ability to proliferate and differentiate into target tissues play a crucial role in novel regeneration methods. These cells are also harvested from various non-ocular sources such as bone marrow, adipose and umbilical cord as well as ocular sources in the corneal layers. Direct and indirect transfer of stem cells with different ocular and ocular references to the cornea's damaged layers and subsequent differentiation into specific corneal cells and its regeneration has been considered a practical treatment in recent years. Due to the high progress in this field in many countries and its replacement with high-risk and time-consuming surgical methods, it needs to be studied more. In this review, in addition to introducing the challenges of current medical procedures and the structure of corneal tissue, the preclinical and clinical advances in the use of stem cells for cell therapy are highlighted.

Synthetic organophosphates (OPs) inhibit acetylcholinesterase resulting in the accumulation of acetylcholine, failure of organs, and eventually death. Diisopropyl-fluorophosphatase (DFPase) is one of the OPs degrading enzymes that has broad substrate from OPs. In this study, for the first time, the secretory expression of DFPase in Bacillus subtilis was investigated in order to accelerate the biodegradation rate of OPs.

DFPase gene was amplified using polymerase chain reaction (PCR) from the pET28-inaV/N-dfpase plasmid. The PCR product was subcloned in the pWB980 plasmid. Competent B. subtilis WB600 were transformed with recombinant plasmid. SDS PAGE technique was used to study the expression of protein secreted in superrich medium.

Appearance of the 946 bp band in agarose gel after digestion of transformed plasmid confirmed the presence of DFPase gene in this construct. Approximately, 35 kDa protein band was shown in culture medium after incubating at 35°C for 72 hours and 150 rpm. Measurement of enzyme’s activity was done by monitoring the release of fluoride from diisopropyl fluorophosphate (DFP), using ion-meter. Results showed that enzyme’s activity was 3333 U/L.

Bacillus subtilis is a suitable host for production of secretory and active form of DFPase.

To evaluate the repeatability of biomechanical readings by Corvis ST (Wetzlar, Germany) in healthy eyes and its relation with age and sex.

Three consecutive measurements were performed on 100 eyes of 100 patients using the Corvis ST. Various parameters including first and second applanation length, first and second applanation velocity, first and second applanation time, peak distance, radius, deformation amplitude, corrected and noncorrected intraocular pressure (IOP), central corneal thickness (CCT), Corvis ST biomechanical index (CBI), and tomography and biomechanical index (TBI) were derived from the Corvis ST readings. Repeatability of each parameter and their correlation with age and sex were evaluated.

The mean IOP, CCT, CBI and TBI were 15.47 ± 2.24 mm Hg, 541.24 ± 38.90 μm, 0.048 ± 0.13 and 0.138 ± 0.17 respectively. Intra class correlation coefficient (ICC) for CCT, second applanation velocity and deformation amplitude were 0.985, 0.809 and 0.825 respectively. ICC was less than 0.8 for all other parameters. The device-specific readings showed no significant relationship with age and sex.

The Corvis ST showed high repeatability for CCT, second applanation velocity, and deformation amplitude parameters. No relation between Corvis ST readings and age or sex was observed.

During the eight years of the imposed war against Iran, various chemical weapons were used by Iraq's Baathist army, including blister (mustard) and nerve gas (Tabun and Sarin).

This study is a review and uses reliable domestic and international sources available through computer databases.

Mustard was the most widely used gas, and more than 30 years after the end of the war, chemical warfare victims still suffer from its harmful effects. Mustard gas is a powerful alkylating agent. The detrimental effects of this gas on different tissues have been reported and its greatest effects on the lungs, eyes and skin have been studied. In this paper, the most important acute and chronic diseases caused by mustard gas in the respiratory system, eyes and skin have been studied. Also, a brief reference was made to nerve gases. Finally, another gas probably used on the battles of the axis of resistance (chlorine gas) was mentioned.

Several types of late lung, eyes, skin, and other complications occurred due to compounds used in deadly chemical weapons, over the years since the end of the Holy Defense and continue to offend the injured. The exact mechanism is unknown.

Organophosphorus hydrolase(OPH) is an enzyme that can degrade organophosphorus compounds in pesticides. High expression levels of OPH in Escherichia coli lead to form inclusion body in cytoplasmic space which is an inactive form of protein and needs a solubilizing and refolding process. The aim of this study was to compare different methods for solubilization and refolding of recombinant OPH expressed in E. coli. OPH was expressed in E. coli and purified by the Ni-NTA column. The refolding efficiency of this protein was assessed by 4 strategies: dialysis, rapid dilution, on column and combination of rapid dilution and dialysis. In each case, the refolding efficiency was evaluated by SDS-PAGE analysis and enzyme activity assay and was compared to find the best procedure. The refolding efficiency of these 4 strategies was estimated at about 12%, 10%, 14% and 50% for on column, rapid dilution, dialysis and combination of rapid dilution and dialysis, respectively. Results showed that during the refolding process, most proteins did not reach the correct structure and aggregated again while the combination of 2 methods, rapid dilution and dialysis provided an appropriate procedure to refold. The combination of rapid dilution and dialysis is an efficient method for refolding OPH. The efficacy of this method for refolding other recombinant proteins can be further investigated.

Macular corneal dystrophy (MCD) is an autosomal recessive hereditary disease. In most cases, various mutations in carbohydrate sulfotransferase 6 (CHST6) gene are the main cause of MCD. These mutations lead to a defect in keratan sulfate synthesis. Retinitis pigmentosa (RP) is another eye disorder with nyctalopia as its common symptom. It has been shown that more than 65 genes have been implicated in different forms of RP. Herein, we report on a 9-member family with 2 girls and 5 boys. Both parents, one of the girls and one of the boys had normal eye vision and another boy had keratoconus. Other children (1 girl and 2 boys) suffered from both MCD and RP. Corneal transplantation and medical supplements were used for MCD and RP during the follow-up period, respectively. Based on the family tree, it seems that the inheritance of both diseases is autosomal recessive. Based on our search of databases, there is no report on the simultaneous presence of MCD and RP. To the best of our knowledge, the present article is the first case report on this topic. Molecular genetic investigation is needed to clarify the mechanism of concurrent MCD and RP.

The aim of the present study was to evaluate the clinical outcomes after implantation of MyoRing in patients with ectasia secondary to LASIK. Patients and

This study was a retrospective, consecutive, nonrandomized interventional case series. The MyoRing was implanted after creation of a stromal pocket using a PocketMaker microkeratome (Dioptex, GmBH, Linz, Austria) in 6 eyes of 6 patients with ectasia secondary to LASIK. Uncorrected distance visual acuity, corrected distance visual acuity, sphere, cylinder and keratometric changes were reported after a 3 year follow-up period.

Uncorrected distance visual acuity and corrected distance visual acuity were improved in 5 and 3 patients respectively. One patient showed decreased UDVA after 3 years and in 3 patients the corrected distance visual acuity decreased at the last visit compared to the preoperative reading. Maximum keratometry, sphere and cylinder were improved from preoperative values in 4, 2 and 5 patients respectively.

Because of the mixed results in our small group of patients, it seems that MyoRing implantation using mechanical dissection is not a very effective method for treatment of patients with post LASIK ectasia. However, large comparative multicenter studies are recommended to further verify these results.Keywords: Ectasia, Cornea, LASIK, Corneal ring, Iran.

The aim of this study was to evaluate the effect of mechanical implantation of MyoRing in keratoconic patients with high myopia.
This study comprised 32 eyes of 32 keratoconic patients (14 males, 18 females) with high myopia. All underwent MyoRing (Dioptex GmbH) implantation using mechanical dissection with a Pocket Maker Microkeratome and completed three months of follow-up. The main outcome measures of the study were preoperative and postoperative uncorrected distance visual acuity (UDVA), corrected distance visual acuity (CDVA), manifest refraction, and keratometry readings.
The mean age of patients was 29.6 ± 6.7years (range 20–44 years). A significant diference was seen in the spherical equivalent refractive error when compared final postoperative examination to preoperative one (-10.51 ± 2.81 D to -1.32 ± 2.29 D) (P˂0.001). The mean UDVA (log MAR) improved significantly from 1.14 ± 0.32 to 0.35 ± 0.24 (P˂0.001) and the mean CDVA (log MAR) improved from 0.47 ± 0.20 to 0.22 ± 0.15 (P˂0.001) postoperatively. The mean of sphere and cylinder of manifest refraction was reduced significantly by 7.70 and 2.6 diopters (D) respectively (p

Marine environment has unique diversity. More than half of biodiversity lives at sea and oceans. Promotion of science and technology led to the future accession to marine species and study them and finally using marine product in human life. Using marine product for treatment of human malady is one of the most attractive applications of this component. Since cancer is the second mortality agent in human and current drugs has several side effects, identification and isolation of anti-cancer component from marine source is one of the most attractive research branches for scientists. The aim of this study is investigation of marine components which used in cancer treatment. In this article, we point to some of anti- cancer and drugs with FDA approve or component that are in different stage of clinical trials. The source of them and their mechanism were investigated too. Despite of an existing variety of fresh and brine source in Iran, there isn’t any spacious study for identification, isolation, investigation and confirmation of remedial, and especially anti-cancer property of marine source. Thus attend to marine source in order to cancer treatment is very necessary.

Observing the correct principles of sterilization is one of the most important
strategies to prevent infection transmission and, also, to reduce the time and the cost of research projects in biological sciences. The aim of this study is to evaluate the awareness of employees in a research laboratory in an institute of medical sciences. In this cross-sectional study, a questionnaire was designed, which
contained important items of sterilization, such as autoclave performance and its quality control, conditions of sterilization by oven, and the principles of surface disinfection, and distributed among the staff. Forty-four filled questionnaires were
collected and the results were analyzed using SPSS version 17. There was a significant correlation between the level of education and the level of researcher's knowledge about the principles of sterilization (P

The use of organophosphorus hydrolase (OPH) enzyme to degrade Chemical Warfare Agents is one of the most frequently used decontamination methods. OPH is a ~36 kDa homodimeric metalloprotein that is found in the membrane of Flavobacterium sp. strain ATCC 27551 and Brevundimonas diminuta MG and is capable of hydrolyzing a wide range of oxon and thion , such as paraoxon and parathion. OPH gene (opd) has been expressed in many hosts, such as bacteria, insect cells, fungi, and Streptomyces spp. High level and soluble expression and correct folding of each protein are of important factors. Fusion proteins, including TRX, Gb1 and MBP, are commonly used to increase solubility, folding and in some cases, stability. The present study evaluated thioredoxin (TRX) role in OPH expression level, solubility and stability by cloning the opd gene into pET32a and pET21a and expressing the resulting vectors in E. coli shuffle T7. The pET32a vector encodes a fusion protein containing TRX that is not present in the pET21a. The results revealed an increased expression level, solubility and stability in OPH produced by the pET32a-opd construct compared to the pET21a vector due to the presence of the TRX fusion in pET32a vector.

Staphylococcus aureus is a hazard to human health since they can cause a wide variety of hospital-associated infections ranging from minor skin infections to post-operative wound infections and food poisoning which produces many different virulence factors, including enterotoxins (SEs). Although studies have been done regarding the difference between virulence factors of sensitive strains and resistant to antibiotics, the aim of this study was to investigate this topic in different patients. This cross-sectional study was performed on 100 patients admitted to a hospital in Tehran. After preparing of wounds samples, antibiogram study was done by disc diffusion method and prevalence of staphylococcal enterotoxin type B or seb gene was confirmed by Polymerase Chain Reaction. Data was analyzed using SPSS 17 software. Results showed that the highest percentage of isolates with positive seb gene is about 7% which related to amoxicillin (6.6%), penicilline and cotrimoxazole (6.5%). More than 90% of isolate are resistant to amoxicillin, penicillin and cotrimoxazol. According to results, a significant relationship between seb gene and resistance to related antibiotics was not observed.

In present study we determine the Zinc and Lead levels in human blood samples by differential pulse polarography under optimum experimental conditions (pH=7.5, Scan rate=5mV/S and pulse amplitude=50mV) the polarographic reduction peaks of Zn and Pb were examinedin the -0.72 V and-0.15 respectively. The zinc and lead content can be determined by in comparison with their standard solutions. According to the obtained results differential pulse polarography is more reliable, rapid and sensitive method for determination of zinc and lead levels in the blood than atomic absorption spectroscopy.

Aim and High efficient transformation of external DNA into the bacterial cells accounts as the main step of microbial biotechnology. For this purpose, some conventional methods have been developed. Using cold CaCl2 is one of the most important and cheapest ways for DNA transformation into the gram negative bacteria such as Escherichia coli. Commonly used methods have a low transformation rate about 105 to 107 transformants per 1μg of DNA. Due to the importance of DNA transformation procedure, numerous attempts have been applied to optimize the conventional methods such as cold CaCl2. In this study we showed that it is possible to increase DNA transformation efficiency into the E. coli in standard CaCl2 method, using a membrane permeable cationic peptide (CM11). Based on ClCa2 competent cell preparation method, the bacterial cells were treated by different concentration of peptide (0.5, 1, 2, 3 and 6 μg/ml) in two different ways. Thereafter pET-28a (+), pGEX4T-1 and pUC19 as model plasmids were separately transformed into the competent cells. Results showed that the highest plasmid transformation efficiency obtained at the present of 1 μg/ml of peptide. In this concentration, the transformation efficiency of pET-28a (+), pGEX4T-1 and pUC19 plasmids were respectively 4.4, 4.7 and 4 fold higher than control sample. The results of this study revealed that in the chemical cold CaCl2 strategy for DNA transformation, CM11 as a cell membrane permeable peptide can increase the efficiency of DNA transformation into E. coli.

Some of bacterial species are able to uptake DNA molecule from environment، the yield of this process depends on some conditions such as plasmid size and host type. In the case of Bacillus subtilis، DNA uptake has low efficacy. Using Spizizen minimal medium is common method in plasmid transformation into B. subtilis، but rate of this process is not suitable and noteworthy. The aim of this study was investigation of novel method for improvement of DNA transformation into B. subtilis based on CM11 cationic peptide as a membrane permeable agent.

In this study، for optimization of pWB980 plasmid transformation into B. subtilis، the CM11 cationic peptide was used. For this purpose، B. subtilis competent cell preparation in the present of different concentration of peptide was implemented by two methods. In the first method، after treatment of bacteria with different amount of peptide for 14h، plasmid was added. In the second method، several concentration of peptide with plasmid was exposed to bacteria simultaneously. Bacteria that uptake DNA were screened on LB agar medium containing kanamycin. The total transformed bacteria per microgram of DNA was calculated and compared with the control.

Plasmid transformation in best conditions was 6. 5 folds higher than the control. This result was statistically significant (P value <0. 001). Discussion and

This study showed that CM11 cationic peptide as a membrane permeable agent was able to increase plasmid transformation rate into B. subtilis. This property was useful for resolution of low transformation efficacy.

Aim and Lipases are the valuable biocatalysts which have many applications in the industry. It is widely used in catalyzing chemical reactions such as water, alcoholic, acidic hydrolysis and also esterification. Bacterial lipases are the most important kinds of enzymes in the industry because they have special advantages such as the high stability in organic solvent, the ability of soft catalyzation of hydrolytic reactions, facility in production process and it’s relatively low cost. Bacillus pumilus is one of the most considerable bacterial sources of this enzyme but the main problem for producing this enzyme is low expression and secretion. The aim of this research is the production of recombinant lipase from native strain in secreting manner for increasing the level of expression in Bacillus subtilis. Lipase-coding gene of bacillus pumilus was cloned into pWB980 plasmid by genetic engineering. After this step, construction plasmids were transferred to Bacillus subtilis WB600 as a host. Then expression and secretion level between native and recombinant forms was evaluated and compared. Secretory form of lipase enzyme was expressed in Bacillus subtilis. An expression level of this enzyme is higher than normal strain. Recombinant lipase production in Bacillus subtilis is a suitable method for increasing the expression of this enzyme.