The primer design of domain-specific C-terminus of the avian H5N1 NS gene from Indonesia

Kindi Adam, Vivi Setiawaty

The primer design of domain-specific C-terminus of the avian H5N1 NS gene from Indonesia

Abstract


Abstrak
Latar belakang: Virus avian influenza (AI) H5N1 telah menyebabkan kematian hampir 600 orang di dunia sejak tahun 2003. Virus AI H5N1 masih beredar pada unggas liar dan domestik. Salah satu sifat
patogenitas virus AI  diketahui dari kemampuan protein nonstructural (NS1 dan NS2) dalam menginduksi badai sitokin. Mutasi yang terjadi pada protein NS1 dan NS2 dapat memperberat infeksi yang terjadi. Metode: Sekuens genetik gen NS yang mengkode domain C-terminus dari virus AI H5N1 didapat dengan metode sekuensing menggunakan primer khusus untuk mengamplifikasi gen NS. Desain primer dilakukan dengan menggunakan beberapa software bioinformatika yang tersedia kemudian primer diuji kembali untuk mengetahui keberhasilan dalam mengamplifikasi sekuen target. Pengujian RT-PCR dilakukan pada
empat isolat virus AI H5N1, dilanjutkan dengan sekuensing untuk mengetahui urutan basa nukleotida yang dihasilkan dari primer yang dibuat dalam penelitian ini. Hasil: Primer yang didesain dapat mengamplifikasi empat isolat H5N1 ayam yang diuji yaitu dua isolat
dari Bali (A/Ck/Bali-KH/2009 dan A/Ck/Bali-T/2009),serta dua isolat dari Jawa Timur (A/Ck/Jawa Timur-S/2009 dan A/Ck/Jawa Timur-D/2009).
Kesimpulan: Primer yang didesain dapat digunakan dalam amplifikasi PCR dan sekuensing gen NS yang mengekspresikan bagian C terminus dari protein NS1 dan bagian ujung NS2. (Health Science Indones
2014;2:83-8)

Kata kunci: avian influenza, H5N1, gen NS, C-terminus

Abstract
Background: The avian influenza (AI) H5N1 viruses have caused nearly 600 fatal cases in the world since 2003. The H5N1 AI virus is still circulating in wild birds and domestic. One of the known of AI virus pathogenicity is the ability of the nonstructural protein (NS1 and NS2) to induce cytokine storm. Mutations of the NS1 and NS2 proteins may aggravate infections.
Methods: Genetic sequences of NS gene that encodes the C - terminus domain of AI H5N1 virus were obtained by sequencing method using specific primer to amplify the NS gene. The primers design were
done by using several bioinformatics software available then retested to determine the primer success in amplifying the target sequence. RT- PCR testing performed on four H5N1 AI virus isolates, followed by sequencing to determine the nucleotide sequences generated from the primer were prepared in this study. Results: Primers, which can be designed to amplify four chicken H5N1 isolates tested were two isolates from Bali (A/Ck/Bali-KH/2009 and A/Ck/Bali-T/2009), and two isolates from East Java (A/Ck/East Java-S/2009 and A/Ck/East Java-D/2009). Conclusion: The designated primer can be used in PCR amplification and sequencing of genes that express NS section C terminus of the protein NS1 and NS2 end portion. The primer design of H5N1 NS gene. (Health Science Indones 2014;2:83-8)

Keywords: avian influenza, H5N1, NS gene, C-terminus


Keywords


avian influenza, H5N1, NS gene, C-terminus

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