z. sharifi

HTLV-1 is the only known oncogenic human retrovirus, linked to ATL and HAM/TSP. Serological methods like ELISA and Western blot have sensitivity limitations, whereas PCR-based techniques offer more reliable detection. The HBZ gene, consistently expressed in ATL and asymptomatic carriers, correlates with proviral load. This study aimed to design specific primers targeting HBZ for a highly sensitive semi-nested PCR assay.

In a Cross-Sectional study, the HBZ gene sequence was identified using the NCBI GenBank database. Sequences from various viral isolates were aligned using MEGA6 bioinformatics software. Primers were designed from a conserved region shared by all HTLV-1 strains. Primer design and evaluation were conducted using Primer-BLAST, Primer3, OLIGO Analyze, SnapGene, and Gene Runner software. For the experimental phase, genomic DNA extracted from blood samples underwent a two-step semi-nested PCR. The PCR products were then analyzed using agarose gel electrophoresis.

The primers designed for the HBZ gene successfully enabled the development of a semi-nested PCR assay. In silico predictions of primer performance were validated after synthesis and laboratory testing, with results aligning perfectly with the bioinformatics predictions.

  This study demonstrates that the primers designed for the HBZ gene, combined with the semi-nested PCR method, provide an effective means for detecting the HTLV-1 provirus. Both the bioinformatics and laboratory results confirmed the specificity and sensitivity of the method, underscoring its potential for rapid and accurate HTLV-1 detection.

During chronic hepatitis B virus infection, the dynamic balance between virus replication and the host immune response is crucial in developing liver disease. In this study, the level of expression of programmed death type 1 in asymptomatic donors infected with hepatitis B during the natural course of the infection was investigated.

In this case-control study, 120 blood donors were divided into two groups. The first control group included 60 healthy individuals who tested negative for HBsAg and anti-HBc. The second group, known as the case group, comprised 60 asymptomatic HBV-infected blood donors who tested positive for HBsAg and anti-HBc. All participants were entered from the Tehran Blood Center in 2018. To check gene expression after RNA extraction and cDNA synthesis, a Real-Time PCR test was performed for the PD-1 gene and β-actin reference gene, then using REST software, gene expression changes compared to the healthy group were checked.

All the samples of asymptomatic HBV-infected blood donors were 100% positive for total anti-HBc, anti-HBe, and negative for HBe Ag. The gene expression levels were analyzed using REST software, utilizing Cycle Threshold  values obtained from Real-Time PCR tests for the PD-1 gene and a reference gene. The analysis revealed 1.5 times increase in PD-1 gene expression in asymptomatic HBV-infected donors compared to healthy donors. However, this increase was not found to be statistically significant.

Conclusions  :

PD-1 gene expression does not change during the normal course of chronic HBV infection in asymptomatic HBV-infected donors and is similar to healthy individuals.

Human lymphotropic virus type I mainly infects lymphocytes. For the detection of this virus, mainly nucleated cells are evaluated. Due to the short longevity of cells, sample collection is a challenge. The purpose of the study was to investigate the presence of HTLV-1 virus in genomic DNA released from cells in plasma and to determine its viral load in Western blot-positive blood donors.

In this descriptive study, 30 HTLV-1 positive Western blot donors were evaluated. Using an extraction kit, genomic DNA was extracted from plasma and isolated PBMC using Ficol. A nested PCR test was used to confirm the presence of HTLV-1 virus. A real-time PCR quantitative test by Sybergreen method was used to determine the amount of virus load.

The average age of subjects was 42.9 ± 13.5 years; 93.3% of them were male and 6.6% female. LTR gene region was detected by nested PCR method in 80% of plasma and 100% of PBMC samples. The median viral loads in plasma and PBMC were 1.90 Copies/µL and 20 copies/106 PBMC, respectively.

Conclusions  :

To detect the HTLV-1 virus by molecular method, the HTLV-1 provirus  DNA released from cells, in the plasma can be used when peripheral blood mononuclear cells are unavailable. It has also been observed that the virus load in the plasma of asymptomatic carriers is approximately 10 times lower than in PBMC.

Self-medication has various implications for children's health. Given the limited documentation on the prevalence of self-medication in children in Iran, this study aims to investigate the frequency of non-academic treatments in children and the phenomenon of self-medication by parents

A cross-sectional study involving 300 parents of hospitalized children at the emergency department of Akbar Children's Hospital in Mashhad was conducted to examine the prevalence of self-medication before hospitalization. A census sampling method was employed. Data were collected using the "Self-Medication and its Prevalence" questionnaire. The collected data were analyzed using SPSS version 23.

The study included 300 children with an average age of 6.09 ± 3.13 years, consisting of 150 boys and 150 girls. Two hundred and eighty-three participants lived in urban areas, and 280 had insurance coverage. Parental self-prescription of medication occurred in 197 cases, with 208 having a history of previous self-prescribed medication. The most common illnesses leading to self-medication in the last 6 months were cold (94.7%), headache (78.7%), and fever (60.7%). The most commonly used medications were cold remedies (93.7%), analgesics (89%), and antibiotics (52.7%)

The findings of this study indicate that self-medication is reported in over 69% of the participants. Education and implementation of various strategies to reduce this phenomenon are recommended, considering its potential consequences.

Various mutations can occur in the HBV genome during chronic HBV infection due to error-prone viral replication and stress on the host immune system. This study aimed to determine the mutations in the BCP and preC regions of the hepatitis B virus genome in asymptomatic carriers and its role in the persistence of hepatitis B infection.

In this descriptive study conducted by easy non-probability sampling method in 2017, 40 asymptomatic blood donors positive in terms of HBsAg and confirmatory tests and negative in terms of anti-HCV and anti-HIV tests were included in the study. Anti-HBc, HBe Ag and anti-HBe (Total) tests and ALT and AST tests were performed. Nested-PCR reaction was performed on the extracted DNA and the sequences were determined. Data analysis was done using online software https://hbv.geno2pheno.org.

Out of the 40 examined samples (94.8%), 38 were men and 2 (5.2%) women with an average age of 38.6 ± 8.6. Mutations in (G1764A) and BCP (A1762T) were observed in 10 (50%) and 8 (40%) cases, respectively, and G54T17 mutation was observed in 3 (15%) samples out of 20 HBV positive and HBeAg negative samples. The mutation in the preC region and the nucleotide position A1896 G was found in 12 cases (60%).

Conclusions  :

The highest frequency of mutations was observed in G1764A and BCP which can be considered as one of the effective factors in not clearing and removing the virus by the host's immune system and causing chronic hepatitis B infection.

Intravenous immunoglobulin (IVIG) is a biological product containing a mixture of IgGs which is needed to protect special patients against various microbial pathogens. Due to worldwide decrease in the prevalence of viral hepatitis, the protective effect of immunoglobulins in treatment needs to be re-examined. For this reason the presence of, anti-HAV, anti-HBs, Anti HBc, anti-HGV, anti-HDV and anti-HEV antibodies in the IVIG products of Iranian plasma have been investigated.

In this randomized descriptive cross-sectional study, the presence or absence of antibodies against hepatitis A, hepatitis E, hepatitis D, hepatitis G, antibodies against hepatitis B surface antigen and hepatitis B core antigen in 38 different lots of IVIG products from the Iranian recovered and source plasma was determined by ELISA method and anti-HAV and anti-HBs antibodies were titrated.

Total anti-HAV in IVIGs was 38304 ± 17735 mIU/mL and this titer was significantly higher in IVIGs from recovered plasma compared with source plasma. Total anti-HBs titer in IVIGs was equal to 2487 ± 965 mIU/mL, and no significant difference was observed between the two types of recovered and source plasma. Anti-HBc, Anti-HEV and Anti-HGV were detected in all products, while no anti-HDV was found.

Conclusions  :

Despite the worldwide decrease in the prevalence of viral hepatitis and reports of a decrease in anti-HAV and anti-HBs titers in IVIG products, IVIG products prepared from Iranian plasma have acceptable anti-HAV and anti-HBs titers according to the European Pharmacopeia and are suitable for replacement therapy.

Nigella sativa L. is a valuable medicinal plant that is widely used in different industries. Accumulation of compatible osmolytes is one of the common responses of plants under drought stress. To investigate the effects of irrigation regimes and biochar (resulting from the heating of cattle manure) on N. sativa, a factorial experiment was conducted in a completely randomized design with three replications in the greenhouse of the Faculty of Agriculture at the University of Kurdistan in 2018. The experimental factors consisted of three drought stress levels (40, 70, and 100% of FC) and two biochar use levels (0 and 15 tons.ha-1). The ANOVA results showed that the interaction effects of drought stress and biochar were significant on hydrogen peroxide, malondialdehyde, superoxide dismutase, peroxidase, proline, soluble carbohydrates (water and ethanol soluble), and osmotic potential. Increasing the intensity of drought stress enhanced the amount of hydrogen peroxide, malondialdehyde, superoxide dismutase, peroxidase, proline, and soluble carbohydrates (water and ethanol soluble) and caused the osmotic potential to become more negative. Biochar application decreased the negative effects of drought stress so that hydrogen peroxide, malondialdehyde, superoxide dismutase, peroxidase, proline, and soluble carbohydrates (water and ethanol soluble) amounts were lower than the treatments without biochar. Overall, the present research results proved the useful and effective role of biochar in improving the physiological traits and protective osmolytes of N. sativa under drought stress.

Alloimmunization is caused by exposure of B lymphocytes to foreign antigens following incompatible blood transfusions and is associated with many hemolytic complications. One of the ways to detect blood group antigens is monoclonal antibodies. The aim of the present study is transforming of alloimmunized cells against KELL antigen in thalassemia patients by EBV-Transformation method and to produce lymphoblastoid cell line producing anti-KELL antibody.

In an experimental study, peripheral blood mononuclear cells (PBMCs) of 10 thalassemia patients immunized against KELL antigen were isolated by ficoll hypaque method and exposed to EBV-containing supernatant of B95-8 cell line for 4 hours. After one to three weeks, the production of antibody-producing lymphoblastoid clones and the specificity of the antibody were examined by microscopic observation and microhemagglutination method, respectively.

The results showed that exposure of PBMCs of thalassemia patients immunized against KELL antigen with EBV viral soup caused the formation of lymphoblastoid clones. Examination of the supernatant of culture of produced clones showed that the antibody produced by these cells were able to identify red blood cells containing KELL antigen and agglutinate them by micro hemagglutination test.

Based on the findings, there is a possibility of PBMC transformation in thalassemia patients immunized against KELL antigen and antibody production.

One of the applications of Epstein-Barr virus in research is the immortalization of B lymphocytes and the creation of the lymphoblastoid cell line. People with thalassemia usually need blood transfusion, the main complication of which is alloimmunization. Antibody against RhE antigen causes hemolytic reactions. The aim of this study is to create a lymphoblastoid cell line that produces anti-E using peripheral blood mononuclear cells of alloimmune thalassemia patients against E antigen when exposed to Epstein Bar Virus (EBV).

In an experimental study, the mononuclear cells of 3 patients were isolated by Ficol and mixed with EBV. The transformed cells were cultured in RPMI medium containing cyclosporine. After 2-3 weeks, the formation of clones in the wells was checked and the hemagglutination test was performed for the wells containing the growing cells; the total amount of antibody in positive wells was measured by ELISA method.

Exposure of mononuclear cells of patients with EBV led to cell transformation and creation of clones in some wells. The results of hemagglutination and ELISA methods showed that the clones were able to produce antibodies against the RhE antigen.

Anti-E antibody-producing lymphoblastoid cell line can be created by using sensitized cells of thalassemia patients against RhE antigen and its immortalization by EBV transformation method.

Human T-cell lymphotropic virus type 1 (HTLV-1) requires cell-to-cell communication to cause infection. The increase in the number of cells infected with virus in turn increases the likelihood of virus transmission and progression of disease. Host immune system responses and genetics factors can affect the susceptibility to virus. Thus, in this study, for the first time the polymorphism in PD-1 gene promoter, which is involved in the negative regulation of immune responses, was evaluated in the HTLV-1 asymptomatic carriers and the healthy controls in the Iranian population in 2020.

In this case-control study, genomic DNAs of 81 HTLV-1 asymptomatic carrier  blood donors and 162 healthy blood donors in Khorasan Razavi province were extracted and the rs36084323 polymorphisms  were genotyped with PCR- RFLP method. To confirm the genotyping results, PCR products were sequenced. The results were analyzed using SPSS-22 statistical software based on chi-square test.

The frequencies of G and A alleles of rs36084323 polymorphism were 75% and 25% in the control group and 79.6% and 20.4% in the case group, respectively. Considering the G allele as a reference there was no significant difference between the case and control groups (p = 0.256).

The results of this study showed that there is no association between G/A in single nucleotide polymorphism of rs36084323 with susceptibility to HTLV-1 virus infection in the Iranian population.

Many‌ people prefer ‌village eggs because of their good taste, ‌bold yolk, and golden skin than commercial eggs, and some people are more interested in commercial eggs because of their production and expiration date codes, and the question always arises which one has a more desirable quality, so this research is aimed at comparing qualitative, microbial and oxidative parameters in the village and commercial eggs in Semnan. In this descriptive cross-sectional study, 75 village and 75 commercial egg samples were collected by random sampling method from retail stores of Semnan and transferred to food hygiene laboratory under cold and sterile conditions to determine qualitative parameters (Haugh unit, egg weight, yolk index, pH, yolk color) and also microbial and oxidative parameters. Village eggs in qualitative parameters except for weight had higher rates compared to commercial eggs (P< 0.05) and in a* and L* color index there was no significant difference in the village and commercial samples. In terms of microbial and oxidative parameters, the comparison of averages showed that village eggs were significantly higher in the overall shell-level and internal egg mesophilic bacterial counts, as well as the village eggs, showed a higher amount of lipid peroxidation compared to commercial ones (P< 0.05). The findings of this study showed that village eggs accounted for more in terms of bacterial counts and the rate of oxidative breakdown, but due to yellow color index and viscosity, they are more marketable than commercial eggs.

Hepatitis B virus (HBV) infection remains a serious global health challenge. Hepatitis B infection is one of the major diseases transmitted by blood transfusion. Infection with the virus can cause extensive liver damage.  So far, 10 genotypes have been identified for hepatitis B. The aim of this study was to do Hepatitis B virus genotyping in asymptomatic HBV blood donors in Tehran.

In this cross-sectional study, 20 HBsAg positive blood donors tested by ELISA were examined. After viral DNA extraction, the pres gene was amplified by Nested-PCR method and the virus genotype was aligned with the references sequence by Clustal w software.

Samples were sequenced after Nested-PCR was done. Hepatitis B virus genotype was determined in all 20 blood donors with hepatitis B to be genotype D. 

Prevalence of genotype D in study subjects was 100%.  This result can be used in preparation of hepatitis B diagnostic kits. Identifying genotypes can also help physicians identify patients at higher risk for disease progression and adopt better treatment strategies.

Human T-cell lymphotropic virus type I (HTLV-I) has been associated with two diseases in humans such as adult T-cell leukemia/lymphoma (ATLL) and HTLV-associated myelopathy/tropical spastic Para paresis (HAM/TSP). About 5 to 10 million people are infected with HTLV-1 worldwide. In order to improve the blood safety and to prevent false positive results in blood donors, methods for identifying infectious agents need to be highly sensitive and specific. The purpose of this study was to compare the electrochemical luminescence, western blotting, and PCR assays to confirm HTLV-1 virus.

This exprimental study was carried out in 2018 on 66 samples (62 males and 4 females) which had antibody against HTLV-1 virus.  All retested samples with the same ELISA kit were examined by electrochemical luminescence and western blotting. For virus detection by PCR method, blood donors were recalled for new samples. After DNA extraction, nested PCR was performed with primers in both TAX and LTR regions.

In this study 8 (%12/1) samples from 66 samples were reacted with ELISA test. Also 4 (%6) samples had antibody against HTLV-1 by electrochemiluminescence test. Finally, four samples were confirmed by Western blotting and Nested PCR.

Conclusions :

The use of the electrochemical luminescence, western blotting and PCR assays, which had similar results in this study, is suitable for detection and confirmation of HTLV-1 virus.

Use of kits with high sensitivity and specificity to determine the prevalence of antibodies against hepatitis B virus (anti-HBc) core antigen is important. The aim of this study was to determine the prevalence of hepatitis B virus antibody in HBsAg negative blood donors using two anti-HBc enzyme immunoassay kits (DIA.PRO and SIEMENS). 

In this Cross-sectional study, a number of 4313 samples of HBsAg negative blood donors were collected. All samples were tested for the detection of anti-HBc (Total, IgM & IgG) by ELISA using Diapro and Siemens kits. Anti-HBc assay was performed by both kits in duplicate. Statistical analysis of data was performed using SPSS Ver. 22.0; chi-square tests were also performed. 

The study population consisted of 227 (5.3%) women and 4086 (94.7%) men. 498 (11.5%) and 384 (8.9%) samples reacted with Diapro and Siemens kits for Anti-HBc, respectively. All positive samples of both kits in duplicate test showed the same results as the previously reported results. 114 cases of reactive kits of DIA.PRO kit were negative in re-testing with SIEMENS kit. Significant differences were observed between the reactive results of the two kits (p = 0.00005).  The kappa coefficient for the reactive results of the two kits was 85%. 

Conclusions:

 Considering the significant difference between the results of the two kits, it is recommended to perform anti-HBc testing with anti-HBs and /or anti-HBe to determine the prevalence of anti-HBc in blood donors with past HBV infection.

In this study, nanoscale zero-valent copper (nZVC) as catalyst activated persulfate (PS) was used for the degradation of p-chlorophenol (p-CP) under ultrasonic (US) irradiation in aqueous solution. The effect of different operational parameters such as solution pH (3.5-10.5), PS concentration (1-7.5 mm/L), nZVC dosage (5-35 mg/L) and initial p-CP concentration (10-100 mg/L) were evaluated at different contact time. Results indicated that US/PS/nZVC system achieve higher efficiency in p-CP degradation than US/PS, US/nZVC and PS/nZVC systems. The optimal p-CP removal efficiency (98%) was achieved within 40 min with 5 mm/L PS and 30mg/L nanocatalyst at 25 mg/L initial p-CP concentration. It was also observed that the p-CP degradation rate depends on initial p-CP concentrations. To clarify the mineralization of p-CP, TOC and COD were analyzed at optimum conditions. COD and TOC removal rate obtained from the US/PS/nZVC system with contact time of 60 min were 61and 75%, respectively. Through the use of methanol (MA) and tert-butyl alcohol (TBA) as radical scavengers,  was identified as the main radical species that are generated during processes. The removal process of p-CP could be described by the pseudo-first-order kinetics. The apparent degradation rate constant (k) was 0.076 min-1 in US/PS/nZVC system at optimal conditions.

Hepatitis C (HCV) infection is one of the main causes of chronic hepatitis diseases all over the world. HCV is a transfusion transmitted virus and a serious threat to general health. HCV genotyping has an important role in tracing routes of infection. This study aimed at investigating the changes in  distribution pattern of HCV genotypes among Iranian blood donors.

In this cross sectional study, 239 HCV confirmed blood donors from Jun. 2015 to Nov. 2017 were included. Semi nested PCR method was used for amplifying the NS5B  region of HCV genome. PCR products were sequenced and HCVgenotypes were determined by searching the sequences in Basic Local Alignment Search Tool (BLAST). Phylogenetic tree was constructed to confirm HCV genotypes. STATA 13 software was used for data analysis. 

In 106 (44.35% ± 6.3%) out of 239 participants, HCV genotype was determined and confirmed. In phylogenetic tree, studied sequences formed three separated clusters. Genotype 3a, 1a and 1b were the common genotypes.

Conclusions :

It seems that molecular epidemiology of HCV infection did not change based on variability of genotypes but changes in the frequency of genotypes have been occurred as a result of replacement of genotype 1a by genotype 3a during the last decade among Iranian blood donors.

Considering the importance of donor selection process in blood safety, this study aimed at comparison of frequency of intravenous drug use (IDU) among accepted HCV positive blood donors in different regions of Iran.

This cross sectional-analytical study was conducted on  serologically confirmed HCV positive blood donors who referred to blood transfusion centers for counseling all over the country during November 2015 to May 2017. All provinces were classified in three regions. IDU status and demographic specification of participants were analyzed using questionnaires. Chi square, fisher exact and ANOVA with STATA software version 13 were used for statistical analysis.

Participants, according to the number of provinces, sample size and geographical proximity to three areas were classified including  76 from North; 105 from West-South and 90 from East- Center individuals with the mean ages of 37.73 ±  8.27, 38.83 ± 8.80 and 36.86 ± 8.63 and  history of IDU of 37 (48.68%), 45 (42.86%) and 41 (45.56%), respectively. No significant differences between different regions of the country were found in the frequency of IDU voluntary blood donors with hepatitis C, age, gender, education status, marriage status (p > 0.05).

Conclusions:

Approximately half of the voluntary blood donors with hepatitis C were IDUs. The frequency of IDU was similar among donors from different regions of Iran. Considering the fact that studied donors were found eligible in donor selection and donated the blood, though rejected during the screening phase, conducting research on the issue can increase the quality of blood donor selection and blood safety.

One of the most important concepts in mathematics, which has always been difficult for students to understand, is the concept of limit. Due to the connection of this concept to many other concepts such as infinitely large and infinitesimally small, continuity, derivative and integral, its correct understanding and comprehension is of particular importance and this has led to its teaching and learning by math educators. Although this concept has been explored many times in educational research by researchers, it is still difficult for students to understand. There are several ways to identify problems in understanding concepts, including the concept of limit. One of these methods is to study how concepts and structures are formed that students create to learn concepts in their minds. The aim of this study is to assess students, understanding of the concept of limit in the third year of secondary school based on the APOS theory. APOS theory is a theory of learning that is used in academic mathematics. The theory categories students’ understanding of concept across the levels, and is able to models mental structures that person to understand of the concept.

 Method and Materials: 

This research is a descriptive study using survey. The sample of this research is 234 students in third grade from Qarchak city who have been randomly selected. The instrument is a researcher-made questionnaire with six questions. The reliability of the test was estimated by Cronbach’s alpha and is approved in the amount of 0.82.

The results show that most students do not have a good understanding of the concept of limit and they mostly can do the limit problems correctly, if they have a routine way to solve them. The weak structures affect not only their understanding of the concept, but also depend on the understanding of the concepts such as continuity.

When introducing the concept of limit, the teacher can prevent the construction of correct schemas of the concept of limit by using slang and giving the initial idea. Because the role of the teacher in constructing a concept of limit is very important, if the teacher teaches in an inappropriate way, it may prevent the student from absorbing the concept of limit. Another reason for stopping the growth and development of the concept is the continuous evaluations in educational environments that do not emphasize the need for conceptual understanding of the concepts and are limited to routine methods to achieve better results. For this reason, doing research in the field of teaching and learning any concept in mathematics, such as the concept of limit, can lead to more effective teaching strategies.

Leishmaniasis is one of the important endemic diseases in Iran and is divided into cutaneous and visceral Leishmaniasis. Since determining the type of the parasite is effective in the controlling and preventing of the disease، we sought to find a method for the diagnosis of cutaneous leishmaniasis.

In this research 35 cutaneous leishmaniasis patients from different part of Iran [8 infectious cases from the province of Isfe-han and its margins namely، Ardestan، Kashan and Samiram; 2 cases from Kerman (the foci of the infection was Sirjan)، 4 cases from Khorasan، 2 cases from Mashhad، 2 cases from Esfraeian، 2 casese from Boshehr، 3 cases from Semnan (the foci of the infection was Damghan) and 1 case from ghom] were collected. After preparing the direct slide DNA extracted from all patients، polymerase chain reaction (PCR) was done for the gene of ITS1.

The results of PCR indicated that all of the cases were positive and showed DNA fragment bands in the size range of 69-350 bp. The results from restriction fragment length of polymorphism (RFLP) indicated that 94% (33 persons) and 6% (2 persons) of the cases were detected as Leishmania major and Leishmania tropica، respectively. Discussion &

These results showed that PCR-RFLP method by using Apo1gene is suitable for the diagnosis of cutaneous leishmaniasis caused by leishmania species.

The purpose of this study was investigate the amount of realization of technical characterization and the assessment of course in developing and designing virtual courses of Iranian universities from the view points of students. Developing and designing virtual courses of criteria was five domain, and because of there are a lot of subjects in this article so, only above categories have been considered. In this study, descriptive-measurable method has been used. Statistical population of the study who were studying graduate and undergraduate students in the courses of engineering and human science who were selected of Shiraz and Khajenasir universities in the colligate year of 1386-87. For gathering the data, were used: quantitative method (researcher-made questionnaire). The questionnaire results identified that, in two research domain (technical characterization, and the assessment of course) The T which was seen was larger than the critical level of table (p≤0.05). In general, the all of the means, was higher than value means (3) (the mean of technical characterization domain 3/58, and the mean of assessment of course domain 3/02). Therefore, in relation to the results of this research as viewed by students, characteristics that mentioned above must pay attention higher average level.

Abstract

Toxoplasmosis is a one of the most world-wide spread zoonosis representing a very serious clinical and veterinary problem. Toxoplasma gondii is an intracellular parasite that causes severe neurologic and ocular disease in immune compromised and congenitally infected individuals. The heavy incidence and severe or lethal damages of toxoplasmosis clearly indicate the need for the development of a more effective vaccine human vaccines are not available and current anti-toxoplasma treatment is disappointing. Immunization with plasmid DNA، a relatively novel technique، is a promising vaccination technique. To improve the immune response by DNA vaccination various is one of the key for the success of the vaccine in the field. One of the most efficient ways to control this disease is immunization. However، so far، there is no effective vaccine available against this pathogen. An important source of human contamination with T. gondii is the consumption of raw or undercooked meat products. Toxoplasma gondii are widely prevalent in humans and other animals which can cause severe or lethal toxoplasmosis. So، the development of a more effective vaccine is needed urgently. Therefore، we prepare gra5 plasmid to use as a vaccine.

In this study، GRA5was cloned in pTZ57R; afterwards، it was transformed into TOP10 strain of E. coli Bacteria. The recombined plasmid extracted from E. coli bacteria and amplified through PCR technique. Besides، pcDNA3 Plasmid for receiving and cloning of GRA5 segment was digested by Hind3 & EcoRI enzymes. GRA5 was sub-cloned into pcDNA3 and the reaction ligation product was transformed for the above bacteria. The bacteria grew in LB culture with ampicillin. Recombined pcGRA5 plasmids were purified from E. coli by Plasmid extraction kit. Finally، recombinant plasmid using cell culture method was expressed in Cho cell.

The accuracy of the results was confirmed by using restriction enzymes and PCR methods. GRA5 was cloned into expression eukaryotic plasmid pcDNA3. After sequencing pcGRA5 plasmid for cell expression Western blot method was used. Discussion &

The results showed that cloning and transformation of fragment GRA5 in pcDNA3 was done properly.