Isolating Barley (Hordeum vulgare L.) B1 Hordein Gene Promoter and Using Sequencing Analaysis For The Identification of Conserved Regulatory Elements By Bioinformatic Tools

Gene expression is a complex multi-step process. For the efficient expression of foreign genes in plants, it is essential to optimize every step of the process for the plant machinery, which includes choosing suitable promoters. Promoters play the most important role in determining the temporal and spatial expression pattern and transcript level of a gene. Some strong constitutive promoters, such as cauliflower mosaic virus 35s promoter, are widely used in plant genetic engineering research. However, the expression levels of the foreign genes in all tissues are often unsatisfied, but some of such problems can be may solved by using a strong seed-specific promoter to restrict gene expression to the seed only. The aim of this article was to characterize a B1 hordein-specific promoter. The promoter region of B1 hordein gene was isolated from the genomic DNA of Walfajre and Alger barley by polymerase chain reaction. Sequence analysis showed that the cloned fragment B hordein promoter (BHP) - contained motifs like TATA box, (CA)n box, ACGT motif, AAAG motif, GCN4-like motif and Ebox, which constituted the seed-specific promoter activity. It was therefore concluded that B1 hordein promoters can be used to engineer and subsequently study stable seed specific gene expression in barley, and potentially to modify barley seeds through genetic engineering.Keywords: Barley, Seed Specific Promoter, Motif, HordeinAfrican Journal of Biotechnology Vol. 11(29), pp. 7378-7387, 10 April, 201

<span style="font-size:11.0pt;font-family: "Times New Roman","serif";mso-fareast-font-family:"Times New Roman";mso-bidi-font-family: Mangal;mso-ansi-language:EN-GB;mso-fareast-language:EN-US;mso-bidi-language: HI" lang="EN-GB">Functional fusion expression of sunflower multicystatin in <i>E. coli</i> and its comparison with a single domain cystatin</span>

375-379<span style="font-size:11.0pt;font-family: " times="" new="" roman","serif";mso-fareast-font-family:"times="" roman";mso-bidi-font-family:="" mangal;mso-ansi-language:en-gb;mso-fareast-language:en-us;mso-bidi-language:="" hi"="" lang="EN-GB">Identification of the molecular structure and novel biophysiological functions of plant cystatins or phytocystatins is of great interest in the field of molecular biology. The important requirements for these are the efficient production, purification and correctly folded forms of these proteins. We report here the cloning, easy expression and characterization of a sunflower multicystatin (SMC) as a functional fusion protein in E. coli. For the first time, the amplified cystatin coding region was expressed as a part of maltose-binding fusion protein using pMALc2X over-expression vector in TB1 strain of E. coli without affecting the recombinant bacterial growth. In comparison to the previously prepared recombinant SMC (rSMC), a high amount (~44 mg/L of bacterial cell culture) of purified fused SMC (fSMC) was obtained using single-step purification method. fSMC strongly inhibited papain activity in vitro as compared to Celosia single-domain cystatin. Purified fSMC may be used for basic biochemical, pharmacological or clinical studies without the cleavage of its fusion parts.</span

Carborundum-dependent entrance of<i> Eco</i>RI restriction enzyme into plant cells and specific cleavage of genomic DNA

684-689In a basic research to determine the morpho-molecular interactions of plant tissues with EcoRI DNA restriction enzyme, it was demonstrated that this protein is capable of entering the sunflower and maize leaf cells using a plant tissue-abrading material and cleaving the genomic DNA at specific sites. This was inferred from the analysis of morphological patterns of EcoRI-treated leaf areas as well as using some molecular tests, including the cleavage pattern analysis of genomic DNA isolated from treated locations followed by ligation of cleaved fragments into EcoRI site of a DNA cloning vector system. The overall results indicated that the specific restriction of genomic DNA may happen following the entrance of EcoRI protein most likely into the nucleus of plant cells

Introduction of an Efficient Multiepitopic Vaccine Against Different SARS-CoV-2 Strains: Reverse Vaccinology

Introduction: In recent years, COVID-19 has been recognized as a health threat. Despite vaccination, people still get the disease because the new variants have mutations in their genomes that allow them to bind to host receptors and evade the immune system’s responses. Therefore, the main aim of this study was to use bioinformatics tools to introduce a rapid and practical vaccine to fight against these diverse mutations in different SARS-CoV-2 strains. Method: To epitopes mapping, 32 different spike protein variants were retrieved. We then used the Immune Epitope Database (IEDB), NetCTL, and NetMHCIIpan to predict T and B cell epitopes. The vaccine based on protected epitopes was evaluated in terms of antigenicity, allergenicity, toxicity, solubility, physicochemical properties, population coverage, and secondary structure with relevant servers. Modeling using Robetta and docking with Toll-like receptor (TLR3) were performed using Cluspro, PatchDock, and FireDock, respectively. Results: After detailed evaluations, all the results confirmed the optimal quality of the vaccine. According to further investigations, this structure is similar to native proteins and there is a stable and strong interaction between the vaccine and the receptor. Based on molecular dynamics simulation, structural compactness and stability in binding were also observed. In addition, the immune simulation showed that the vaccine can stimulate immune responses similar to real conditions. Finally, codon optimization and in silico cloning confirmed efficient expression in Escherichia coli. Conclusion: Based on the obtained results, the designed multi-epitope vaccine can serve as a prophylactic candidate against SARS-CoV-2

Effects of Human Erythropoietin on Functional Outcome of Patients with Traumatic Cervical Cord Injury; A Pilot Randomized Clinical Trial

Objective: To determine the effects of recombinant human erythropoietin (rhEPO) on functional outcome and disability of patients with traumatic cervical spinal cord injury (SCI). Methods: This was a randomized, double blind, placebo controlled clinical trial being performed in Nemazee and Shahid Rajaei hospitals of Shiraz during a 3-year period from 2011 to 2014. A total number of 20 patients with acute traumatic cervical SCI less than 8 hours after injury were included. We excluded those with anatomic cord dissection, penetrating cord injury and significant concomitant injury. Patients were randomly assigned to receive rhEPO in 500IU/mL dosage immediately and 24-hour later (n=11) or placebo (n=9). All the patient received standard regimen of methylprednisolone. Neurological function was assessed on admission, 1, 6 and 12 months after the injury according to the American Spinal Cord Injury Association (ASIA). Results: Overall we include a total number of 20 patients. The mean age of the patients was found to be 40.1±9.5 (ranging from 19 to 59) years. There were 18 (90.0%) men and 2 (10.0%) women among the patients. There was no significant difference between two study groups regarding the baseline characteristics. The baseline ASIA score was comparable between two study groups. The motor and sensory ASIA scores were comparable between two study groups after 1, 6 and 12 months follow-ups. We also found that there was no significant difference between two study groups regarding the motor and sensory outcome in complete cord injury and incomplete cord injury subgroups. Conclusion: Administration of rhEPO does not improve the functional outcome of patients with traumatic cervical SCI. Clinical trial registration: The study has been registered with Iranian Registry for Clinical Trials (www.irct. ir; IRCT2014122920471N1

Anaesthetic Efficacy of 4% Articaine in Comparison with 2% Lidocaine as Intraligamentary Injections after an Ineffective Inferior Alveolar Nerve Block in Mandibular Molars with Irreversible Pulpitis: A Prospective Randomised Triple-Blind Clinical Trial

The objective of the current study was to compare the anaesthetic efficacy of supplemental intraligamentary (IL) injection of 4% articaine with that of 2% lidocaine in the mandibular first and second molars with irreversible pulpitis after an ineffective inferior alveolar nerve block injection (IANB) using the same anaesthetic in a randomised triple-blind clinical trial. Seventy-six adult patients, who were diagnosed with irreversible pulpitis in the mandibular first or second molars, were divided into 2 groups and received IANB randomly. In patients with lip numbness, anaesthesia was evaluated with the cold and electrical pulp (EPT) tests, and if the reported number on EPT was below 100, supplemental IL injection was administered using the same anaesthetic. The teeth were retested after 5 minutes. The Heft–Parker visual analogue scale was used to evaluate pain after IANB and IL injections. Statistical analysis was performed using repeated measures ANOVA, chi-square, and independent-sample and paired-sample t-tests. The results showed that there was no significant difference in the success rates of supplemental IL and IANB injections between articaine and lidocaine. Furthermore, there was no significant difference in the success rates of supplemental IL injection with lidocaine between the mandibular first and second molars. However, there was a significant difference in the success rates of supplemental IL injection with articaine between the mandibular first and second molars. Moreover, supplemental IL injections indicated no significant difference in the anaesthetic efficacy between articaine and lidocaine; nevertheless, they were more effective in the mandibular second molars, especially with articaine