Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1
Avian Influenza viruses belonging to the Orthomyxoviridae family are enveloped viruses with segmented negative sense RNA genome surrounded by a helical symmetry capsid. Influenza viruses, especially the highly pathogenic avian influenza virus (HPAI) such as H5 or H7 subtype are the most important pa...
| Main Authors: | , , , |
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| Format: | Conference or Workshop Item |
| Language: | English |
| Published: |
2009
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| Online Access: | http://psasir.upm.edu.my/id/eprint/65127/1/PGM-1-7.pdf |
| _version_ | 1825932886322184192 |
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| author | Abubakar, Mustapha Bala Ideris, Aini Omar, Abdul Rahman Bejo, Mohd Hair |
| author_facet | Abubakar, Mustapha Bala Ideris, Aini Omar, Abdul Rahman Bejo, Mohd Hair |
| author_sort | Abubakar, Mustapha Bala |
| collection | UPM |
| description | Avian Influenza viruses belonging to the Orthomyxoviridae family are enveloped viruses with segmented negative sense RNA genome surrounded by a helical symmetry capsid. Influenza viruses, especially the highly pathogenic avian influenza virus (HPAI) such as H5 or H7 subtype are the most important pathogens for the poultry industry in recent times. The haemagglutinin protein and neuraminidase, serves as the target for the immune response of the host. Due to recurrent genetic reassortments between avian and human influenza viruses, global pandemics may emerge and the naive human immunity could not withstand pressure by the novel hybrid virus. The emergence of genetic engineering technology provided the industry with new methods of manufacturing diagnostics tools and vaccines. After extraction of RNA from the cell culture of strain influenza A/Chicken/Malaysia/2004(H5N1) of AIV, the viral RNA was converted to cDNA by a specific primer. The cDNA was amplified by the polymerase chain reaction (PCR) and analyzed by agarose gel electrophoresis. The intact PCR product of full length haemagglutinin gene was cloned in TOPOTM TA Cloning vector. The full-length HA-encoding gene of H5N1 AIV was subcloned into a pPICZA vector. After successful ligation, the constructed plasmid was transformed into E.coli.Top10, Plasmid DNA from transformed bacteria was extracted in white colony and positive clones were confirmed by restriction digestion with Sacl and Not1 restriction enzymes, colony PCR screening and nucleotide sequencing. Construction of a recombinant pPICZA/H5HA plasmid containing the full length haemagglutinin gene was achieved as a first step towards the expression in Pichia pastoris. |
| first_indexed | 2024-03-06T09:48:50Z |
| format | Conference or Workshop Item |
| id | upm.eprints-65127 |
| institution | Universiti Putra Malaysia |
| language | English |
| last_indexed | 2024-03-06T09:48:50Z |
| publishDate | 2009 |
| record_format | dspace |
| spelling | upm.eprints-651272018-09-04T04:04:09Z http://psasir.upm.edu.my/id/eprint/65127/ Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 Abubakar, Mustapha Bala Ideris, Aini Omar, Abdul Rahman Bejo, Mohd Hair Avian Influenza viruses belonging to the Orthomyxoviridae family are enveloped viruses with segmented negative sense RNA genome surrounded by a helical symmetry capsid. Influenza viruses, especially the highly pathogenic avian influenza virus (HPAI) such as H5 or H7 subtype are the most important pathogens for the poultry industry in recent times. The haemagglutinin protein and neuraminidase, serves as the target for the immune response of the host. Due to recurrent genetic reassortments between avian and human influenza viruses, global pandemics may emerge and the naive human immunity could not withstand pressure by the novel hybrid virus. The emergence of genetic engineering technology provided the industry with new methods of manufacturing diagnostics tools and vaccines. After extraction of RNA from the cell culture of strain influenza A/Chicken/Malaysia/2004(H5N1) of AIV, the viral RNA was converted to cDNA by a specific primer. The cDNA was amplified by the polymerase chain reaction (PCR) and analyzed by agarose gel electrophoresis. The intact PCR product of full length haemagglutinin gene was cloned in TOPOTM TA Cloning vector. The full-length HA-encoding gene of H5N1 AIV was subcloned into a pPICZA vector. After successful ligation, the constructed plasmid was transformed into E.coli.Top10, Plasmid DNA from transformed bacteria was extracted in white colony and positive clones were confirmed by restriction digestion with Sacl and Not1 restriction enzymes, colony PCR screening and nucleotide sequencing. Construction of a recombinant pPICZA/H5HA plasmid containing the full length haemagglutinin gene was achieved as a first step towards the expression in Pichia pastoris. 2009 Conference or Workshop Item PeerReviewed text en http://psasir.upm.edu.my/id/eprint/65127/1/PGM-1-7.pdf Abubakar, Mustapha Bala and Ideris, Aini and Omar, Abdul Rahman and Bejo, Mohd Hair (2009) Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1. In: 8th Malaysia Congress on Genetics, 4-6 Aug. 2009, Genting Highlands, Malaysia. (pp. 180-184). |
| spellingShingle | Abubakar, Mustapha Bala Ideris, Aini Omar, Abdul Rahman Bejo, Mohd Hair Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 |
| title | Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 |
| title_full | Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 |
| title_fullStr | Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 |
| title_full_unstemmed | Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 |
| title_short | Molecular cloning of full-length haemagglutinin (H5HA) gene of highly pathogenic avian influenza virus (HPAI) H5N1 |
| title_sort | molecular cloning of full length haemagglutinin h5ha gene of highly pathogenic avian influenza virus hpai h5n1 |
| url | http://psasir.upm.edu.my/id/eprint/65127/1/PGM-1-7.pdf |
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