BAB 5 KESIMPULAN DAN SARAN
5.2 Saran
Protein rekombinan human IFN α2a pada penelitian masih perlu dilakukan pemisahan fusi tag dan purifikasi sehingga didapatkan protein rekombinan yang murni. Protein rekombinan human IFN α2a yang telah murni perlu diuji aktivitasnya secara in vitro mengunakan sel mamalia dan uji in vivo menggunakan hewan uji dengan dibandingkan IFN α2a komersial. Hasil penelitian ini juga perlu dilakukan scale up untuk mendapatkan protein rekombinan human IFN α2a dalam jumlah yang lebih banyak.
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Lampiran 1. Skema Kerja Penelitian
Amplifikasi dengan PCR Pemotongan
Purifikasi Purifikasi fenol kloroform
Pemotongan
Purifikasi fenol kloroform
Skrining dan verifikasi klon transforman
(PCR colony screening, isolasi vektor rekombinan, analisis restriksi, sekuensing)
Kultivasi dan isolasi vektor rekombinan pET-32b(+)-IFN α2a
Skrining dan verifikasi klon transforman (PCR colony screening)
Kultivasi dan ekspresi sel transforman (induksi dengan IPTG)
Isolasi protein rekombinan human IFN α2a Analisis elektroforesis SDS-PAGE (Western blot dan Pewarnaan CBB)
Penyiapan
vektor pET-32b(+) Penyiapan
DNA insert
Ligasi DNA insert dan vektor pET-32b(+)
Transformasi ke E. coli DH5α (inang kloning)
Transformasi ke E. coli BL21(DE3) (inang ekspresi)