Construction of Novel Phytochelatins by Overlap Oligonucleotides

Synthetic phytochelatins are protein analogs of phytochelatin with similar heavy metal binding affinities that can be easily produced from a synthetic DNA template. We design synthetic phytochelatin [(Glu-Cys)n Gly] linked to hexahistidine by viral linker peptide and then followed by gene synthesis and cloning of it. Then peptide coding gene (synthetic phytochelatin with linker and hexahistidine) was designed exactly and constructed with step by step methods by overlapping oligonucleotides using T4 DNA Ligase. Finally, synthesized gene amplified by PCR, cloned in pTZ57R/T and transformed to Escherichia coli (DH5α). The results of sequencing show that some types of synthetic phytochelatin (EC4, EC12, and EC20) with linker and hexahistidine were constructed and cloned in vector

Optimization of expression, purification and handling anti bacteria feature protein of bovine neutrophil B-defensing BNBD2

زمینه و هدف: دیفنسین‎ها یکی از بزرگ‎ترین خانواده‎ی پپتید‎های ضد میکروب می‎باشند که به واسطه‎ی فعالیت بر علیه باکتری‎ها، قارچ‎ها و بسیاری از ویروس‎ها به عنوان آنتی‎بیوتیک‎های نسل جدید منفعت بسیاری دارند. هدف از این مطالعه بهینه سازی بیان، تخلیص و بررسی خاصیت ضد میکروبی پروتئین بتا دیفنسین 2 نوتروفیل‎های گاو (BNBD2) بوده است. روش بررسی: در این مطالعه‎ی تجربی-آزمایشگاهی باکتری اشرشیاکلی B‏L21(DE3) حامل وکتور pET-32a(+) که ژن BNBD2 در آن همسانه سازی شده بود جهت مطالعات مورد استفاده قرار گرفت. بیان پروتئین BNBD2 با تغییر در پارامترهای دانسیته‎ی سلولی، دمای رشد، مدت زمان القاء با استفاده از سیستم الکتروفورز عمودی (SDS-PAGE) و تست برادفورد به صورت کمی و کیفی بررسی گردید. مراحل تخلیص پروتئین نوترکیب با کمک روش شیمیایی شکافت در جایگاه فرمیک اسید و عبور از سانتریکون و اثر ضد باکتری پروتئین تخلیص شده بر چند نمونه‎ی باکتریایی گرم مثبت و گرم منفی مورد بررسی قرار گرفت. یافته ها: با استفاده از محیط کشت Luria–Bertani، شروع القاء در جذب نوری 8/0 در طول موج 600 نانومتر، غلظت یک میلی مولار ماده‎ی القاء کننده‎ی IPTG، دمای رشد 30 درجه و مدت زمان 4 ساعت پس از القاء بیشترین میزان بیان پروتئین به دست آمد. پروتئین نوترکیب با استفاده از شکافت در جایگاه فرمیک اسید و عبور از سانتریکون تخلیص گردید. نتایج آزمایش وسترن بلاتینگ نیز نشان داد که پروتئین نوترکیب به طور اختصاصی به آنتی‎بادی mouse anti-(His)6 peroxidase متصل می‎گردد. تشکیل هاله‎ی عدم رشد در محیط‎های کشت مولر هینتون آگار حاوی کشت سطحی باکتری های مورد آزمایش خاصیت ضد باکتری این پروتئین را نشان داد. نتیجه گیری: با توجه به خاصیت ضد میکروبی پروتئین BNBD2و امکان بیان پروتئین در باکتری E. coli می توان به تولید انبوه این پروتئین نوترکیب اقدام نمود

Toward an integrated sustainable urban design framework in the historic center of Baghdad

The old center in Baghdad is well defined by its unique urban fabric, surrounded by modern urban pattern and by modern roads, which replaced its walls. Narrow alleys, natural shading, mixed use, human scale, privacy, and walkable and natural environments are the most significant features in the historic center, which also represent the main principles of urban sustainability. The historic area in Baghdad is currently suffering from problems such as air pollution, a lack of modern facilities, traffic congestion, uncontrolled land use, and a low standard of infrastructure. These problems have brought into focus the extent to which a sustainable urban design framework can provide appropriate solutions to regenerate the traditional fabric in terms of urban form, land use, transportation, and create a new vision to deal with the social and economic processes. This paper will explore how the traditional urban fabric in Baghdad might be the perfect solution for implementing urban sustainability. Consequently, our challenge in this paper is to find a solution that can serve as a platform to resolve the conflicting values of traditional urban form and modern design models, sourcing new sustainable technologies and the integration of the infrastructure systems

Children as frequent attenders in primary care: a systematic review

Background: Frequent paediatric attendances make up a large proportion of the general practitioner (GP) workload. Currently no systematic reviews on frequent paediatric attendances in primary care exists. Aim: To identify the socio-demographic and clinical characteristics of children who attend primary care frequently. Design and setting: A systematic review.Methods: The electronic databases MEDLINE, EMBASE and PsycINFO were searched up to January 2020, using terms relating to frequent attendance in primary care settings. Studies were eligible if they considered children frequently attending in primary care (0-19 years). Relevant data were extracted and analysed by narrative synthesis.Results: Six studies, of overall fair quality, were included in the review. Frequent attendance was associated with presence of psycho-social and mental health problems, younger age, school absence, presence of chronic conditions, and high level of anxiety in their parents.Conclusions: Various sociodemographic and medical characteristics of children were associated with frequent attendance in primary care. Research on interventions needs to account for the social context and community characteristics. Integrating GP services with mental health and social care could potentially provide a response to medical and psycho-social needs of frequently attending children and their families

Immunoreactivity of various peptides in typical and atypical bronchopulmonary carcinoid tumours.

The presence of a number of regulatory peptides (bombesin, gastrin, glucagon, somatostatin, calcitonin and ACTH) was compared in 30 typical carcinoid tumours and 27 well differentiated neuroendocrine carcinomas (atypical carcinoids) using conventional immunocytochemistry. Strong immunostaining for one or more peptide was observed in 97% of the typical carcinoids (29/30) whereas only 67% of the neuroendocrine carcinomas showed immunoreactivity. The peptide most frequently detected in typical carcinoids was bombesin (67%), while gastrin was more common in neuroendocrine carcinomas (44%). Immunoreactivity for more than one peptide was present in 33 tumours and in three cases, six different peptides were detected. The study shows that immunoreactivity to various peptides is more common in typical carcinoids than well differentiated neuroendocrine carcinomas. The significance of these findings is discussed

Chondroprotective Effect of Zerumbone on Monosodium Iodoacetate Induced Osteoarthritis in Rats

The objective of this investigation was to evaluate chondroprotective effect of zerumbone, a purified compound of Zingiber zerumbet Smith against monosodium iodoacetate (MIA) induced knee osteoarthritis (OA) in the rat. The effect on the articular cartilage was examined and compared with celecoxib (Celebrex®), a Non-Steroidal Anti-Inflammatory Drug (NSAID). Forty adult male Sprague Dawley rats were divided into four groups (n=10 for each). All animals were injected with MIA intraarticularly in their right knee joints to induce OA. Rats from first and second groups were treated with zerumbone in a same dose but with two different concentrations. Rats in the third group were treated with celecoxib and served as positive control whereas the fourth group were treated with corn oil and served as negative control. Evaluation of OA changes in the knees was assessed with the aid of both radiography and histopathology score. Macroscopic as well as microscopic examinations revealed curative effect of zerumbone in a dose dependent manner on the osteoarthritic knee joints. Apart from this, our data also revealed very poor anti-OA property of celecoxib. We concluded that oral administration of zerumbone in a dose of 2 mL kg-1 b.wt. of 0.4% w/v diluted with corn oil for a period of 4 weeks has some chondroprotective effects

Long-term discolouration modelling for cast iron mains

Water companies have been working to introduce strategies to reduce discolouration customer contacts via non-specialist ‘business as usual’ practices. A greater understanding of discolouration material behaviour, however, is still needed to accurately inform the mobilisation response and regeneration rates in mains of different materials. The Variable Condition Discolouration Model (VCDM) that tracks both accumulation and mobilisation processes has been validated in some pipe materials using long-term time series data. This paper investigates calibration for a 15 km cast iron (CI) main, using daily turbidity responses with VCDM parameter sensitivity and temporal stability investigated using a statistical approach comparing three periods of the data. Results highlight the VCDM as widely applicable to determine long-term discolouration behaviour and improve behavioural understanding. In this case, analysis of different time periods indicates flow-conditioning not only improves network resilience but can also reduce mobilisation rates and discolouration risk

Eurotherm Seminar N° 81 Reactive Heat Transfer in Porous Media

Abstract. Combustion in Porous Medi

Utilization of Waste Iron Powder as Fine Aggregate in Cement Mortar

This paper reports about the use of recycled iron powder (IP) in producing cement mortar under normal conditions. Flow table test was performed on fresh mortar. Destructive tests were conducted on cubes of the hardened mortar to obtain the compressive and flexural strengths of the cement mortar. The effects of adding 10%, 20%, 30%, and 40% of waste IP as a natural sand replacement were assessed and compared. Waste iron are of two types: iron IP, which shows a similar particle size distribution to that of the sand used in making the samples, and fine iron powder (FIP), which contains fine particles. The compressive strength decreased with the increased amount of added IP in the mixtures, but it increased with the addition of 10% FIP and decreased gradually with the increased FIP level. By contrast, the flexural strength significantly increased with increased FIP in the mixtures. Recommendations regarding the applications of recycling to conserve resources and raw materials and prevent environmental pollution are provided

A Roadmap for the Production of a GMP-Compatible Cell Bank of Allogeneic Bone Marrow-Derived Clonal Mesenchymal Stromal Cells for Cell Therapy Applications

Background: Allogeneic mesenchymal stromal cells (MSCs) have been used extensively in various clinical trials. Nevertheless, there are concerns about their efficacy, attributed mainly to the heterogeneity of the applied populations. Therefore, producing a consistent population of MSCs is crucial to improve their therapeutic efficacy. This study presents a good manufacturing practice (GMP)-compatible and cost-effective protocol for manufacturing, banking, and lot-release of a homogeneous population of human bone marrow-derived clonal MSCs (cMSCs). Methods: Here, cMSCs were isolated based on the subfractionation culturing method. Afterward, isolated clones that could reproduce up to passage three were stored as the seed stock. To select proliferative clones, we used an innovative, cost-effective screening strategy based on lengthy serial passaging. Finally, the selected clones re-cultured from the seed stock to establish the following four-tired cell banking system: initial, master, working, and end of product cell banks (ICB, MCB, WCB, and EoPCB). Results: Through a rigorous screening strategy, three clones were selected from a total of 21 clones that were stored during the clonal isolation process. The selected clones met the identity, quality, and safety assessments criteria. The validated clones were stored in the four-tiered cell bank system under GMP conditions, and certificates of analysis were provided for the three-individual ready-to-release batches. Finally, a stability study validated the EoPCB, release, and transport process of the frozen final products. Conclusion: Collectively, this study presents a technical and translational overview of a GMP-compatible cMSCs manufacturing technology that could lead to the development of similar products for potential therapeutic applications. Graphical Abstract: [Figure not available: see fulltext.