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Design and Production of a Novel Recombinant Chimeric IL2-Omp31 Antigen against Brucella Infection | ||
| Archives of Razi Institute | ||
| مقاله 4، دوره 73، شماره 3، آذر 2018، صفحه 199-206 اصل مقاله (575.45 K) | ||
| نوع مقاله: Original Articles | ||
| شناسه دیجیتال (DOI): 10.22092/ari.2017.110504.1131 | ||
| نویسندگان | ||
| M. Naghavi؛ M. H. Sekhavati* ؛ M. Tahmoorespur؛ M. R. Nassiri | ||
| Department of Animal Science, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran | ||
| چکیده | ||
| Brucellosis is a zoonotic disease in human and animals. Brucella melitensis is one of the most pathogenic species of Brucella in goat and sheep. Omp31 is an outer membrane protein of Brucella that acts as an immunogenic protein. Cytokines are glycoproteins with low molecular weight that play the role of an immune adjuvant and regulate immune responses. Interleukin-2 is one of the most important cytokines, which are secreted by the white blood cells and involved in T cell immune responses. In the present study, a chimeric Omp31-Interleukin2 recombinant protein was generated by means of genetic engineering techniques. This chimeric coding sequence was amplified by using specific primers and using Splicing Overlap Extension (SOE) PCR technique. The fusion of the two mentioned proteins was accomplished using a rigid linker. The generated chimeric IL2-Omp31 fragment was TA cloned, and then subcloned into pEt22b vector as an expression vector. The chimeric protein was successfully expressed in E. coli BL21 (DE3) and confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and also Western-blotting analysis. Finally, in order to assess the antigenic features of the recombinant chimeric IL2-Opm31 protein, its secondary structure and antigenicity were predicted in silico. | ||
| کلیدواژه‌ها | ||
| Brucella melitensis؛ Interleukin؛ Omp31؛ Cytokines | ||
| عنوان مقاله [English] | ||
| طراحی و تولید یک فیوژن آنتی ژن نوترکیب IL2-Omp31 بر علیه بیماری عفونی بروسلا | ||
| نویسندگان [English] | ||
| مهدیه نقوی؛ محمد هادی سخاوتی؛ مجتبی طهمورث‌پور؛ محمد رضا نصیری | ||
| گروه علوم دامی، دانشکده کشاورزی، دانشگاه فردوسی مشهد، مشهد، ایران | ||
| چکیده [English] | ||
| بروسلوز یک بیماری مشترک بین انسان و دام محسوب می شود. یکی از سویه های پاتوژن این بیماری در گوسفند و بز بروسلا ملیتنسیس می باشد. Omp31 یک پروتئین خارج غشائی از باکتری بروسلا بوده و نقش مهمی در تحریک سیستم ایمنی میزبان ایفا می کند. سیتوکین ها گلیکوپروتئن های با وزن مولکولی کم می باشند که نقش بسزائی در ایمنی میزبان و همچنین به عنوان تقویت کننده سیستم ایمنی میزبان بشمار می آیند. اینترلوکین 2 یکی از مهمترین سیتوکین ها در تحریک ایمنی سلولی می باشد و توسط سلول های خونی سفید تولید و ترشح می شود. در این مطالعه با استفاده از تکنیک های مهندسی ژنتیک یک فیوژن آنتی ژن نوترکیب از دو پروتیئن Omp31 و اینترلوکین 2 با هدف تولید واکسن نوترکیب استفاده گردید. توالی کد کننده این فیوژن آنتی ژن به وسیله روش SOE-PCR تکثیر گردید. در این تحقیق، به منظور اتصال بین دو پروتئین از لینکر سخت استفاده شد. توالی کد کننده فیوژن آنتی ژن در وکتور TA کلون گردید و سپس در به وکتور بیانی pET22b منتقل گردید. پروتئین فیوژ شده با موفقیت در سویه بیانی اشرشیا کلی BL21 (DE3) بیان و کیفیت پروتئین تولید شده توسط روش SDS-PAGE و وسترن بلات مورد ارزیابی قرار گرفت. پروتیئن مورد نظر با وزن مولکولی 46 کیلو دالتون مشاهده گردید. در نهایت، در محیط in silico شکل فضائی فیوژن آنتی ژن تولید شده و همچنین قدرت آنتی ژنیسیتی آن مورد بررسی و مطالعه قرار گرفت. | ||
| کلیدواژه‌ها [English] | ||
| بروسلا ملیتنسیس, اینترلوکین, Omp31, سیتوکین | ||
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