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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| 格式: | Thesis |
| 语言: | English English |
| 出版: |
2003
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| 主题: | |
| 在线阅读: | http://psasir.upm.edu.my/id/eprint/9582/1/FSAS_2003_41_IR.pdf |
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| 总结: | 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. |
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