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Pengaruh konsentrasi inducer dan waktu overekspresi pada produksi protein rekombinan fim-C Salmonella typhi
Demam tifoid merupakan masalah kesehatan masyarakat di dunia , seperti Indonesia. Salah satu upaya dalam pencegahan kasus demam tifoid yaitu dengan vaksinasi. Vaksin-vaksin yang telah digunakan dalam mengatasi demam tifoid antara lain vaksin Ty21a dan Vi-Polisakarida, akan tetapi vaksin-vaksin tesebut memiliki kelemahan antara lain timbul efek samping seperti mual, demam dan sakit kepala. Saat ini masih terus dikembangkan vaksin rekombinan yang jauh lebih aman dan pada umumnya tidak menimbulkan efek samping. Tujuan penelitian ini untuk mendapatkan informasi tentang pengaruh konsentrasi inducer IPTG (Isopropyl-β-D-thiogalactopyranoside) dan waktu overekspresi pada produksi protein rekombinan Fim-C Salmonella typhi dalam skala laboratorium sebelum diproduksi pada skala yang lebih besar. Proses overekspresi dilakukan dengan penginduksian dengan variasi konsentrasi IPTG yaitu 0.4 mM, 0.5 mM, dan 0.6 mM serta dengan waktu overekspresi yang bervariasi yaitu 4 jam, 5 jam, dan 6 jam pada 50 mL media LB cair untuk mencari kondisi yang optimal. Setelah mendapatkan konsentrasi IPTG dan waktu overekspresi yang optimal dilakukan proses overekspresi pada 100 mL media LB cair. Pengukuran konsentrasi protein rekombinan Fim-C Salmonella typhi dilakukan dengan menggunakan metode BCA (Bicinchoninic Acid). Karakterisasi protein rekombinan Fim-C dilakukan dengan menggunakan metode SDS-PAGE. Hasil karakterisasi SDS-PAGE menunjukkan pita overekspresi protein rekombinan Fim-C Salmonella typhi pada ukuran ± 31 kDa. Berdasarkan data hasil penelitian dapat disimpulkan bahwa konsentrasi IPTG 0.5 mM dan waktu overekspresi 4 jam merupakan kondisi optimum pada produksi protein rekombinan Fim-C Salmonella typhi. Kata Kunci: Salmonella typhi, protein rekombinan Fim-C S. typhi, IPTG (isopropyl-β-D-thiogalactopyranoside), metode SDS-PAGE
Typhoid fever is a public health problem in the world, such as Indonesia. One of the efforts in the prevention of cases of typhoid fever is by vaccination. Vaccines that have been used in overcoming typhoid fever include Ty21a and ViPolysaccharide vaccine, but these vaccines have disadvantages such as side effects such as nausea, fever and headache Currently still developed recombinant vaccine is much safer and generally does not cause side effects. The purpose of this study was to obtain information about the effect of IPTG (Isopropyl-β-Dthiogalactopyranoside) inducer concentration and the time of overexpression on production of recombinant Fim-C protein Salmonella typhi in laboratory scale before being produced on a larger scale. The process of overexpression was done by induction with IPTG concentration variations of 0.4 mM, 0.5 mM, and 0.6 mM and with varying time of overexpression of 4 hours, 5 hours, and 6 hours on 50 mL of liquid LB medium to find optimal conditions. After obtaining IPTG concentration and optimal overexpression time, the process of overexpression was done on 100 mL of liquid LB medium.The measurement of recombinant protein concentration of Fim-C Salmonella typhi was done by using BCA (Bicinchoninic Acid) method. Characterization of Fim-C recombinant protein was performed using the SDS-PAGE method. The result of characterization of SDS-PAGE showed overimpresi recombinant protein band Fim-C Salmonella typhi at size ± 31 kDa. Based on the research data, it can be concluded that 0.5 mM IPTG concentration and 4 hours of overexpression time is the optimum condition in recombinant protein production of Fim-C Salmonella typhi. Keyword: Salmonella typhi, recombinant protein Fim-C S. typhi, IPTG (isopropyl-β-D- thiogalactopyranoside), SDS method
SS00016146 | SK 16146 | UPT Perpustakaan UNJ (CD.03.2018.003) | Tersedia |
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