ENGLISH: Comparative Evaluation of Human Papillomavirus Type 16 L1 Protein Expressed in Plasmid-and Baculovirus-Based Systems in Insect Cells (FRENCH: É valuation Comparative de la Proté ine de Type 16 L1 du Papillomavirus Humain Exprimé e dans des Systè mes à Base de Plasmides et de Baculovirus dans des Cellules d'Insectes)

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ABEDI KIASARI B.

Title

ENGLISH: Comparative Evaluation of Human Papillomavirus Type 16 L1 Protein Expressed in Plasmid-and Baculovirus-Based Systems in Insect Cells (FRENCH: É valuation Comparative de la Proté ine de Type 16 L1 du Papillomavirus Humain Exprimé e dans des Systè mes à Base de Plasmides et de Baculovirus dans des Cellules d'Insectes)

Author(s)

ABEDI KIASARI B. | Issue Writer Certificate

Keywords

ENGLISH:Baculovirus

Cloning

Expression

Papillomavirus (FRENCH:le Baculovirus;le Papillomavirus;l'Expression;le Clonage)

Abstract

ENGLISH: Human papillomavirus (HPV) has been associated with specific types of papillomas, lesions at particular anatomic sites, and malignancies. The HPV16 and HPV18 have been shown to play a role in a variety of carcinomas. The most documented HPV-associated cancer is cervical carcinoma. Suitable antigens are needed to be identified for the diagnostic tests and vaccines and the Expression of L1 recombinant protein should be accelerated in papillomaviruses. Therefore, in this study, the Expression of the L1 protein of HPV16 was evaluated and compared in insect cells using a plasmid and a baculovirus system. The Expression of the L1 protein of HPV16 in insect cells was investigated using a plasmid (InsectDirect) and a baculovirus system (BacMagic). The expressed recombinant proteins were purified from the Sf9 lysate using Ni-NTA resin columns. The characterization of recombinant L1 protein expressed in both systems (BacMagic and InsectDirect) was performed using immunofluorescence, SDS-PAGE, western blot, and dot blot. The yields of the purified proteins from the plasmid-and baculovirus-based systems (10 ml culture; 107 cells) had the ranges of 455-495 μ g/ml and 1. 44-1. 6 mg/ml as analyzed by spectrophotometer, respectively. The SDS-PAGE analysis of purified proteins revealed that the recombinant proteins with the expected size of 58 KDa were produced in both InsectDirect and baculovirus systems. A high degree (95%) of purification was achieved using this system as observed in SDS-PAGE. The purified L1 protein in the baculovirus system was clearly more efficient than the InsectDirect system. The results of this study indicate that the BacMagic system is an appropriate tool for large scale protein production and provides an alternative to the traditional baculovirus system. In addition, the InsectDirect system might provide a rapid and dependable pointer of whether a protein can be successfully produced in a baculovirus system. Both InsectDirect and BacMagic systems present remarkable savings in cost and time. FRENCH: Ré sumé : Le papillomavirus humain a é té associé aux types spé cifiques de papillomes, aux lé sions sur des sites anatomiques particuliers et aux tumeurs malignes. Les HPV16 et HPV18 ont un rô le causal dans une varié té de carcinomes. Le carcinome du col uté rin est le cancer le mieux documenté associé au HPV. Dans cette é tude, l'Expression de papillomavirus de type 16 L1 dans des cellules d'insectes à l'aide d'un plasmide et d'un systè me baculovirus a é té é valué e et comparé e afin de fournir des antigè nes approprié s pour les tests de diagnostic et les vaccins ainsi que pour 'accé lé rer l'Expression de la proté ine recombinante L1 des papillomavirus. L'Expression de la proté ine de type 16 L1 du papillomavirus dans des cellules d'insectes a é té é tudié e en utilisant un plasmide (InsectDirect) et un systè me baculovirus (BacMagic). Les proté ines recombinantes exprimé es ont é té purifié es à partir du lysat Sf9 en utilisant des colonnes de ré sine Ni-NTA. La caracté risation de la proté ine L1 recombinante exprimé e dans les deux systè mes (BacMagic et InsectDirect) a é té ré alisé e par immunofluorescence, SDS-PAGE, Western blot et dot blot. Les rendements des proté ines purifié es analysé s par spectrophotomé trie dans le systè me plasmidique (culture de 10 ml; 107 cellules) allait de 455 μ g / ml à 495 μ g / ml, alors que le systè me à base de baculovirus (culture de 10 ml; 107 cellules) produisait 1, 44 mg / ml à 1, 60 mg / ml de proté ines recombinantes. Une analyse SDS-PAGE des proté ines purifié es a ré vé lé que les proté ines recombinantes de taille attendue (58 KDa) é taient produites dans les systè mes InsectDirect et baculovirus. Un degré é levé de purification (95%) a é té atteint en utilisant ce systè me, comme observé dans le profil SDS-PAGE. La proté ine L1 purifié e dans le systè me baculovirus é tait clairement plus efficace que le systè me InsectDirect. Les ré sultats de cette é tude indiquent que le systè me BacMagic est un systè me approprié pour la production de proté ines à grande é chelle et constitue une alternative au systè me baculovirus traditionnel. Le systè me InsectDirect peut é galement fournir un indicateur rapide et fiable afin d’ é valuer si une proté ine peut ê tre produite avec succè s dans un systè me à baculovirus. Les systè mes InsectDirect et BacMagic pré sentent tous deux des gains de temps et une é conomie de frais remarquable.

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APA: Copy

ABEDI KIASARI, B.. (2020). ENGLISH: Comparative Evaluation of Human Papillomavirus Type 16 L1 Protein Expressed in Plasmid-and Baculovirus-Based Systems in Insect Cells (FRENCH: É valuation Comparative de la Proté ine de Type 16 L1 du Papillomavirus Humain Exprimé e dans des Systè mes à Base de Plasmides et de Baculovirus dans des Cellules d'Insectes). ARCHIVES OF RAZI INSTITUTE, 75(2), 187-195. SID. https://sid.ir/paper/776005/en

Vancouver: Copy

ABEDI KIASARI B.. ENGLISH: Comparative Evaluation of Human Papillomavirus Type 16 L1 Protein Expressed in Plasmid-and Baculovirus-Based Systems in Insect Cells (FRENCH: É valuation Comparative de la Proté ine de Type 16 L1 du Papillomavirus Humain Exprimé e dans des Systè mes à Base de Plasmides et de Baculovirus dans des Cellules d'Insectes). ARCHIVES OF RAZI INSTITUTE[Internet]. 2020;75(2):187-195. Available from: https://sid.ir/paper/776005/en

IEEE: Copy

B. ABEDI KIASARI, “ENGLISH: Comparative Evaluation of Human Papillomavirus Type 16 L1 Protein Expressed in Plasmid-and Baculovirus-Based Systems in Insect Cells (FRENCH: É valuation Comparative de la Proté ine de Type 16 L1 du Papillomavirus Humain Exprimé e dans des Systè mes à Base de Plasmides et de Baculovirus dans des Cellules d'Insectes),” ARCHIVES OF RAZI INSTITUTE, vol. 75, no. 2, pp. 187–195, 2020, [Online]. Available: https://sid.ir/paper/776005/en

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ISSN: 2588-4824

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