DESAIN DNA PELACAK SECARA IN SILICO SEBAGAI PENDETEKSI MUTASI PADA GEN embB Mycobacterium tuberculosis
Abstract
ABSTRAK: Terapi lini pertama Tuberkulosis (TB) selalu menggunakan terapi kombinasi yaitu rifampisin, isoniazid, etambutol, dan pirazinamid. Penggunaan etambutol dalam terapi kombinasi TB digunakan untuk mencegah terjadinya resistensi terhadap obat lain namun tingkat resistensi etambutol secara bertahap meningkat. Mutasi pada operon embCAB bertanggung jawab pada resistensi etambutol dengan prevalensi tertinggi terjadi pada kodon 306 gen embB. Mutasi pada gen embB kodon 306 juga dikaitkan dengan adanya kecenderungan resistensi akibat peningkatan konsumsi obat sehingga dijadikan sebagai kandidat potential marker untuk board drug resistance, khususnya untuk MDR-TB. Penelitian ini akan mendesain urutan nukleotida TaqMan probe untuk mendeteksi mutasi M306I menggunakan program Clone Manager Suite 9.2. Hasil rancangan probe DNA kemudian dianalisis berdasarkan kriteria probe secara umum dan berdasarkan kriteria pelabelan TaqMan probe. Rancangan probe DNA mutan menggunakan program menghasilkan 10 probe yang memenuhi kriteria probe secara umum untuk mutasi M306I pada gen embB. Berdasarkan analisa pelabelan TaqMan probe, diperoleh 7 probe (E306MI4, E306MI5, E306MI6, E306MI7, E306MI9, E306MI10, dan E306MI13) untuk deteksi mutasi M306I pada gen embB. Hasil rancangan probe mutan yang telah memenuhi kriteria pelabelan TaqMan probe dapat digunakan untuk mendeteksi adanya mutasi kodon 306 gen embB Mycobacterium tuberculosis. Hasil perancangan TaqMan probe perlu diuji secara eksperimental untuk membuktikan efisiensi kerja dari probe tersebut agar dapat digunakan pada metode Real-Time PCR.
ABSTRACT: First-line Tuberculosis (TB) therapy always uses combination therapy, such as rifampicin, isoniazid, ethambutol, and pyrazinamide. Ethambutol was effective for preventing treatment failures caused by Mycobacterium tuberculosis isolates resistant to other anti-TB drugs however, the resistance rate of ethambutol has gradually increased. Mutations in the embCAB operon have been identi?ed to confer resistance to ethambutol, with embB codon 306 being the most frequently affected. embB306 mutations are associated a tendency for resistance due to increasing numbers of antibiotics consumption so it may be a potential marker for broad drug resistance, especially for MDR-TB. This research design the TaqMan probe nucleotide sequence for the M306I spesific mutation using the Clone Manager Suite 9.2 program. The results of the DNA probe design were then analyzed based on probe criteria in general and based on the TaqMan probe labeling criteria. The mutant DNA probes design using the program produced 10 probes thats have met the general probe criteria for the M306I mutation in the embB gene. Based on the TaqMan probe labeling analysis, theres 7 probes (E306MI4, E306MI5, E306MI6, E306MI7, E306MI9, E306MI10 and E306MI13) for the detection of M306I mutations in the embB gene. The results of the mutant probe design that has met the TaqMan labeling criteria can be used to detect mutations in M. tuberculosis embB gene codon 306. The results of the TaqMan probe need to be tested experimentally to prove the working efficiency of the probe so that it can be used in the Real-Time PCR method.
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References
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