Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus

The haemagglutinin-neuraminidase (HN) glycoprotein of Newcastle disease virus (NDV) is of primary importance in inducing virus-neutralizing antibodies against viral infection in chicken and has been used in the development of many vaccines. A variant strain of the vaccine strain V4QUE known as V4...

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Main Author:	Tang, Yik Kiong
Format:	Thesis
Language:	English English
Published:	2003
Subjects:	Newcastle disease virus Thermostat
Online Access:	http://psasir.upm.edu.my/id/eprint/9582/1/FSAS_2003_41_IR.pdf
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spelling	my-upm-ir.95822023-11-27T06:51:21Z Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus 2003-07 Tang, Yik Kiong The haemagglutinin-neuraminidase (HN) glycoprotein of Newcastle disease virus (NDV) is of primary importance in inducing virus-neutralizing antibodies against viral infection in chicken and has been used in the development of many vaccines. A variant strain of the vaccine strain V4QUE known as V4UPM(HR) has been developed as a heat stable vaccine for use in the poultry industry in tropical countries such as Malaysia. This protein may also be involved in maintaining heat stability of some vaccine strains. In this study, the HN gene of the heat stable variant NDV strain V4UPM(HR) and its parental strain V4QUE were cloned and expressed in the Baculovirus Expression Vector System (BEVs) and characterized for their heat stability. The 1.9 kb HN genes of these strains were amplified by RT-PCR from their genomic RNA and unidirectionally cloned into the baculovirus transfer plasmid, peR Bac4.8. These recombinant baculovirus plasmids were then co-transfected with linearized baculoviral DNA, Bac-N-Blue™ DNA into Spodoptera frugiperda (Sf9) insect cell line. The recombinant baculoviruses which were generated as recHNV4UPM(HR) and recHNV4QUE, were purified by plaque assay. The respective recombinant HN glycoproteins (recHNs ) which were expressed in Sf9 insect cells showed haemagglutination (RA) and neuraminidase (NA) activities as well as haemagglutination inhibition (HI) and haem adsorption activities in serological assays. The HA and NA activities were also detected on the surface and in the cytoplasm of the infected Sf9 cells. SDS-PAGE and Western blot analysis of the recombinant baculovirus-infected Sf9 cell lysates detected protein bands of approximately ~74 kDa, which corresponded to the glycosylated HN protein of the virion. These results indicated that the recHNs were not only successfully expressed in the S19 cells but they also appeared to be biologically active and functional. Newcastle disease virus Thermostat 2003-07 Thesis http://psasir.upm.edu.my/id/eprint/9582/ http://psasir.upm.edu.my/id/eprint/9582/1/FSAS_2003_41_IR.pdf text en public masters Universiti Putra Malaysia Newcastle disease virus Thermostat Faculty of Science and Environmental Studies Mohd. Yusoff, Khatijah English
institution	Universiti Putra Malaysia
collection	PSAS Institutional Repository
language	English English
advisor	Mohd. Yusoff, Khatijah
topic	Newcastle disease virus Thermostat
spellingShingle	Newcastle disease virus Thermostat Tang, Yik Kiong Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus
description	The haemagglutinin-neuraminidase (HN) glycoprotein of Newcastle disease virus (NDV) is of primary importance in inducing virus-neutralizing antibodies against viral infection in chicken and has been used in the development of many vaccines. A variant strain of the vaccine strain V4QUE known as V4UPM(HR) has been developed as a heat stable vaccine for use in the poultry industry in tropical countries such as Malaysia. This protein may also be involved in maintaining heat stability of some vaccine strains. In this study, the HN gene of the heat stable variant NDV strain V4UPM(HR) and its parental strain V4QUE were cloned and expressed in the Baculovirus Expression Vector System (BEVs) and characterized for their heat stability. The 1.9 kb HN genes of these strains were amplified by RT-PCR from their genomic RNA and unidirectionally cloned into the baculovirus transfer plasmid, peR Bac4.8. These recombinant baculovirus plasmids were then co-transfected with linearized baculoviral DNA, Bac-N-Blue™ DNA into Spodoptera frugiperda (Sf9) insect cell line. The recombinant baculoviruses which were generated as recHNV4UPM(HR) and recHNV4QUE, were purified by plaque assay. The respective recombinant HN glycoproteins (recHNs ) which were expressed in Sf9 insect cells showed haemagglutination (RA) and neuraminidase (NA) activities as well as haemagglutination inhibition (HI) and haem adsorption activities in serological assays. The HA and NA activities were also detected on the surface and in the cytoplasm of the infected Sf9 cells. SDS-PAGE and Western blot analysis of the recombinant baculovirus-infected Sf9 cell lysates detected protein bands of approximately ~74 kDa, which corresponded to the glycosylated HN protein of the virion. These results indicated that the recHNs were not only successfully expressed in the S19 cells but they also appeared to be biologically active and functional.
format	Thesis
qualification_level	Master's degree
author	Tang, Yik Kiong
author_facet	Tang, Yik Kiong
author_sort	Tang, Yik Kiong
title	Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus
title_short	Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus
title_full	Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus
title_fullStr	Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus
title_full_unstemmed	Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus
title_sort	thermostability of the recombinant haemagglutinin-neuraminidase glycoproteins of newcastle disease virus
granting_institution	Universiti Putra Malaysia
granting_department	Faculty of Science and Environmental Studies
publishDate	2003
url	http://psasir.upm.edu.my/id/eprint/9582/1/FSAS_2003_41_IR.pdf
_version_	1794018851618291712

Thermostability of the Recombinant Haemagglutinin-Neuraminidase Glycoproteins of Newcastle Disease Virus

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