نشریه مهندسی ژنتیک و ایمنی زیستی
سال نهم شماره 1 (پیاپی 17، بهار و تابستان 1399)

Cancer treatment using medicinal plants has long been of interest to researchers. In the meantime, some plant species contain substances which can inhibit or eliminate cancer cells through apoptosis or necrosis. The family of Lamiaceae and Asteraceae are among the medicinal plants that have many biological effects. The purpose of this study was to investigate the anticancer effect of Pennyroyal and Artemisia extracts on gastric cancer cell line (AGS). In this study, aerial parts of plants were extracted. AGS cells were treated with different concentrations of hydro-alcoholic extract (50-1000 µg/ml) over 24, 48 and 72 hours. Cell motility was estimated using MTT assay and rate of apoptosis induction was measured by flow cytometry with The Annexin-V. The results of MTT assay indicated strong and concentration dependent prevention of cancer cell proliferation by various concentrations of Pennyroyal and Artemisia extracts. These extracts had a dose and time dependent anticancer effect on AGS cells. By increasing the concentration of the extract and incubation for 72 hours, the highest percentage of cell death was observed. In the study of apoptosis in treated cells, Pennyroyal extract was more effective. Due to the cytotoxic effects of hydro-alcoholic extracts on AGS cells, these plants could be used as potential option for further studies on cancer treatment. Therefore, the purification of the active substances in these extracts and the determination of their effect mechanism are recommended.

Biotin sulfoxide reductase enzyme is one of the enzymes involved in adaptation to different environmental conditions. BisC enzyme converts biotin sulfoxide to biotin and Methionine sulfoxide to Methionine under oxidative stress conditions. Both free Methionine and protein bound chains Methionine are essential in protein synthesis process and the function of proteins. The activity of the BisC enzyme is dependent to the Bis-MGD as a co-factor. In the current study, in order to verify the presence of bacterial Molybdeum cofactor, an indirect approach was made by showing the activity of BisC enzyme as a reporter gene. Therefore, in this regard BisC gene was cloned to pJet vector and was transferred to Mach1 strain of E. coli by heat shock transformation. The clones were selected on LB medium complemented with 200 µg/ml of Ampicillin antibiotic and the PCR reaction was done. After verifying of positive clones by sequencing, the BisC gene was digested and ligated to the pETDuet expression vector and transferred to Rossetta. Over-expression of BisC enzyme was analyzed on a SDS-PAGE gel and the activity of the enzyme could be showed as a colorless band on the Native gel by oxidation of methyl viologen. It proved the presence of active Bis-MGD as a cofactor.

The residual of pesticides in the agricultural production is endangering human health and environmental balance. Concerns about pesticide residues and environmental impacts due to repeated use of pesticides has led to investigations of the residue and fate of these agents. The adverse effects of pesticideschr('39') residue on humans and the environment have increased interest in genetic engineering technology and cultivation of genetically modified crops for pest management strategies. This study was carried out to investigate the residue of permethrin (EC 25%) in a Vendor cultivar of tomato in greenhouse.  Tomato plants were sprayed at doses of 0.5- 1 g/lit. Samples preparation was performed by the QuEChERS method. Further purification was achieved using a silica solid-phase extraction (SPE) cartridge and pesticide residues were analyzed using GC-MS. This study revealed that residue decreased in descending order. Results showed that the Permethrin (0.5, 1 g/lit) levels were detected below than maximum residue level (MRL) recommended by codex (0.5 mg/kg) at 5 and 7 days after application. Permethrin levels (1 g/lit treatment) were detected below MRL 10 days after application.

Bacterial wilt caused by Ralstonia solanacearum is one of the most destructive diseases which negatively affects the tomato production worldwide. Considering the importance of the disease, using biological control agents could be effective approaches in reducing the damage of the pathogen. Plant specimens suspected to be infected by tomato bacterial wilt disease were collected from different regions of East Azerbaijan province in Iran. Phenotypic identification of the strains was performed using recommended biochemical and physiological tests. In genotypic assessment, using PCR, a 282 and 148 bp fragments with the PS96I/ PS96H primers were amplified. Based on phenotypic and genotypic properties, the strains that cause wilt of tomato were identified as R. solanacearum biovar 2. For biological control of this disease, 15 bacterial strains were collected from rhizosphere of healthy tomato plants using soil serial dilution method. Antagonistic effect of the strains was evaluated based on their growth inhibitory on pathogen. According to the results, three bacterial strains showed antagonistic activity against R. solanacearum. Based on phenotypic and genotypic characteristics, the antagonist strains Ka1, Ka2 and Ka3 were identified as Bacillus subtilis and Pseudomonas fluorescens, respectively.

Lactoferrin, which consists of lactoferampin and lactoferricin peptides, is one of the milk glycoproteins with anti-microbial properties. Lactoferrin is known as a component of intrinsic immunity which points to first barrier against microbes. The aim of this study was the chimeric synthesis of this two peptides and also producing transgenic product. One of the desired methods to produce recombinant peptides is by transferring their genes to plant cells and producing hairy roots. In this study, the sequence of chimeric Lfampin-Lfcin was codon optimized based on Tobacco expression system. The target sequence was artificially synthesized in pGH vector and then sub-cloned into the pBI121 expression vector. The expression vector was transferred to Agrobacterium rhizogenes through freez- defreezing process. Then it was transferred to Tobacco plant using leaf disc method. Extraction of recombinant protein was done using Tris-HCl method and purification process was accomplished by chromatography Ni-agarose columns. Finally, the results showed that the amount of recombinant protein was 1.52 mg/ml and its quality was also desirable. The results also showed that chimeric composition has a good inhibitory effect on pathogenic microorganisms.

Silybum marianum L. from the Asteraceae is an annual or biennial plant widely distributed in the north, west and south of Iran. The fruits are containing medicinal flavonoids such as silymarin. This study was conducted to optimize the growing conditions for hairy roots and to assay the diverse factors affecting the induction and growth of hairy roots induced by Agrobacterium rhizogenes. The seeds were cultured on MS medium and were kept in the growth chamber. After germination and seedling growth; explants from the leaves, hypocotyl and roots were taken behind growth and after 5, 10 and 15 days. The explants were inoculated (treated) with two (R1000 and 15834) Agrobacterium rhizogenes strains. Furthermore, the effects of two inoculation methods (floating and injection) were assayed on the hairy roots emergence percentage. MS and 1/2 MS growing media were compared regarding the establishment of the hairy roots in the liquid medium. In a separate experiment, the effect of sucrose amount (20, 30 and 40 gl-1) were evaluated on hairy root induction and the roots emergence percentage was recorded. To verify the transgenic identity, PCR test with specific marker genes was carried out. The results revealed that 15834 strain was the strain of choice in 5 days old hypocotyl explants and produced more hairy roots than other strain. The best rooting percentage belonged to leaf explants at 15 days-old, treated with R1000. Floating was the method of choice for inoculation. 30 gL-1 sucrose was the best treatment for the root induction as well.

Drought is one of the most important environmental stresses for plants, which leads to a significant reduction in agricultural production. In drought stress conditions, plants exhibit a variety of responses at the physiology and sub-cellular levels such as increased expression of some genes like dehydrines. They are categorized in Group 2 proteins Late Embryogenesis Abundant proteins (LEA) that are synthesized and accumulated in different plant tissues not only in response to stress, but also as part of a pre-planned phenomenon during the final stages of development of the seed. Different types of Dehydrine proteins have been identified in Arabidopsis, clover and other plants. Due to the dry and semi-arid climate of Iran and the high economic importance of safflower, in this study, drought stress was applied to this plant with use of methanol spraying in a completely randomized design. The expression of dehydrine 1 in the seed formation stage using the RT-qPCR method was evaluated and the means were compared using SPSS ver. 18. The results showed that the highest level of drought stress (50%) associated with methanol spraying (30%), had significantly the highest level of dehydrine 1 gene expression whereas in other cases the interaction of methanol and drought stress did not have a statistically significant effect on dehydrine 1 expression. It is suggested that methanol spraying can be used as a suitable method for the plant to deal with the effects of stress in low rainfall environments.

The emergence of the dangerous and deadly COVID-19 disease in late 2019 in Wuhan, China, and its rapid spread has had a profound global impact on health, mental security, economy, culture and politics of various countries, drawing the attention of the medical community and related sciences for identification and treatment of this disease. Various studies on this disease have identified a new coronavirus as the cause. Due to the development of severe respiratory infection and the similarity of this virus to SARS-CoV, it was named SARS-CoV-2. Although many studies have been conducted to identify the origin of this virus and based on the obtained results, most researchers have proposed the hypothesis of natural selection of mutation in the bat-RaTGG13, but some researchers also believe that this virus was engineered in a laboratory based on the evidence obtained. However, both groups of researchers believe that none of the hypotheses can be refuted until strong evidence is found as to the origin of the virus. In this review, along with the discussion on the origin of SARS-CoV-2 virus, the introduction of this virus, its function, pathogenesis and ways to prevent and treat the disease will be introduced.