Dvelpoment And Optimization Of Multiplex Pcr For The Detection Of Avian Influenza Strains In Pakistan
By: Mirza Salman Saleem
| Asso. Prof. Dr. Muhammad Hanif.
Contributor(s): Prof. Dr. Khushi Muhammad | Faculty of Veterinary Sciences.
Material type: 
BookPublisher: 2009Subject(s): Department of MicrobiologyDDC classification: 1148,T Dissertation note: The pathogenic Influenza A viruses (subtype H5N1, H7N2 and H9N3), are emerging avian influenza (AI) viruses that have been causing global concern as a potential pandemic threat. Some forms having zoonotic importance (H5N1 and H7N7). So it is a matter of priority to develop quick and efficient methods for detection of Influenza viruses.
For the detection of avian influenza, HA (haemagglutination) test and HI (haemagglutination inhibition) tests are being used for long time. But studies have shown that Influenza virus shows variability and diversity and a high rate of mutation, which makes diagnosis difficult. For this reason the reverse transcriptase PCR (RT-PCR) assays are considered to be a helpful tool.
In this study design, a multiplex RT-PCR strategy was optimized and developed for the detection of AI virus (subtypes H5, H7 and H9). Primers were designed from sequence available Influenza Database (IVDB) for Pakistan and neighboring regions. The primers were annealed at different temperatures so as to optimize a temperature at which all three primers can amplify their respective subtypes. The results clearly indicated that a multiplex RT-PCR is a quick and efficient method for the detection and it is also economical as fewer reagents are utilized. The PCR products of the reaction can potentially be used to provide additional information about strain variation, either by restriction analysis or PCR product sequencing.
The core objectives achieved are the development of an efficient and economical method for detection of avian influenza viruses by designing indigenous primers and optimization of a multiplex RT-PCR for the avian influenza virus.
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Thesis
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UVAS Library Thesis Section | Veterinary Science | 1148,T (Browse shelf) | Available | 1148,T |
The pathogenic Influenza A viruses (subtype H5N1, H7N2 and H9N3), are emerging avian influenza (AI) viruses that have been causing global concern as a potential pandemic threat. Some forms having zoonotic importance (H5N1 and H7N7). So it is a matter of priority to develop quick and efficient methods for detection of Influenza viruses.
For the detection of avian influenza, HA (haemagglutination) test and HI (haemagglutination inhibition) tests are being used for long time. But studies have shown that Influenza virus shows variability and diversity and a high rate of mutation, which makes diagnosis difficult. For this reason the reverse transcriptase PCR (RT-PCR) assays are considered to be a helpful tool.
In this study design, a multiplex RT-PCR strategy was optimized and developed for the detection of AI virus (subtypes H5, H7 and H9). Primers were designed from sequence available Influenza Database (IVDB) for Pakistan and neighboring regions. The primers were annealed at different temperatures so as to optimize a temperature at which all three primers can amplify their respective subtypes. The results clearly indicated that a multiplex RT-PCR is a quick and efficient method for the detection and it is also economical as fewer reagents are utilized. The PCR products of the reaction can potentially be used to provide additional information about strain variation, either by restriction analysis or PCR product sequencing.
The core objectives achieved are the development of an efficient and economical method for detection of avian influenza viruses by designing indigenous primers and optimization of a multiplex RT-PCR for the avian influenza virus.
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