Medical Journal of The Islamic Republic of Iran (MJIRI)
Iran University of Medical Sciences
Fri, Nov 29, 2024
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Volume 32, Issue 1 (2-2018)
Med J Islam Repub Iran 2018 |
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Hosseini S S, Taheri K, Hosseini S M, Gholami M, Kouhsari E, Edalati E, et al . Extraction and purification of the H9N2 virus nucleoprotein: A simple and practical method
. Med J Islam Repub Iran 2018; 32 (1) :754-759
URL: http://mjiri.iums.ac.ir/article-1-5202-en.html
URL: http://mjiri.iums.ac.ir/article-1-5202-en.html
Seyedeh Saeedeh Hosseini 
, Kayvan Taheri , Seyedeh Marzieh Hosseini , Mehrdad Gholami , Ebrahim Kouhsari , Elahe Edalati , Rasoul Madani , Rokhsareh Mohammadzadeh , Abed Zahedi Bialvaei , Mohammad Sholeh , Fariba Golchin Far
, Kayvan Taheri , Seyedeh Marzieh Hosseini , Mehrdad Gholami , Ebrahim Kouhsari , Elahe Edalati , Rasoul Madani , Rokhsareh Mohammadzadeh , Abed Zahedi Bialvaei , Mohammad Sholeh , Fariba Golchin Far
Department of Botany, Biotechnology and Bioinformatics, Payame Noor University, Tehran, Iran , az1171355@gmail.com
Abstract: (3421 Views)
Background: Avian Influenza disease annually entails a significant economic loss to the poultry industry around the world. Influenza virus is a polymorphic virus of the orthomyxoviridae family (single-stranded RNA genome), and nucleoprotein (NP) is the structural and internal protein of the virus. The aim of the work was to purify nucleoprotein for further investigations with a simple, low-cost, fast and practical method.
Methods: In this study, H9N2 influenza virus was isolated in specific pathogen-free embryonated chicken eggs by allantoically inoculating 103 to 105 egg-infective doses (EID50) for 9 to 11 days, purified by 10% (W/V) polyethylene glycol (PEG) 6000 with a sucrose gradient of 60% to 30%. The influenza virus proteins were collected and prepared as fractions by preparative electrophoresis. Finally, the purified NP was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot procedures.
Results: The protein analysis with SDS-PAGE and silver nitrate staining indicated that the desired samples contained purified nucleoprotein and lacked other viral proteins. The results of the investigation of lyophilized fractions containing nucleoprotein on the SDS-PAGE revealed the absence of viral RNA in nucleoprotein and its high purity.
Conclusion: According to this study, purified nucleoprotein can be used to produce nucleoprotein vaccines, as well as to study structural, molecular and diagnostic and therapeutic materials.
Methods: In this study, H9N2 influenza virus was isolated in specific pathogen-free embryonated chicken eggs by allantoically inoculating 103 to 105 egg-infective doses (EID50) for 9 to 11 days, purified by 10% (W/V) polyethylene glycol (PEG) 6000 with a sucrose gradient of 60% to 30%. The influenza virus proteins were collected and prepared as fractions by preparative electrophoresis. Finally, the purified NP was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot procedures.
Results: The protein analysis with SDS-PAGE and silver nitrate staining indicated that the desired samples contained purified nucleoprotein and lacked other viral proteins. The results of the investigation of lyophilized fractions containing nucleoprotein on the SDS-PAGE revealed the absence of viral RNA in nucleoprotein and its high purity.
Conclusion: According to this study, purified nucleoprotein can be used to produce nucleoprotein vaccines, as well as to study structural, molecular and diagnostic and therapeutic materials.
Type of Study: Original Research |
Subject:
Virology
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