Optimasi Konsentrasi MgCl2 dan Suhu Annealing pada Proses Amplifikasi Multifragmens mtDNA dengan Metoda PCR

Mukhammad Asy’ari; A. Saifuddin Noer

doi:10.14710/jksa.8.1.23-27

DOI: https://doi.org/10.14710/jksa.8.1.23-27

Optimasi Konsentrasi MgCl2 dan Suhu Annealing pada Proses Amplifikasi Multifragmens mtDNA dengan Metoda PCR

Mukhammad Asy’ari¹

, A. Saifuddin Noer²

¹Chemistry Department, Faculty of Sciences and Mathematics, Diponegoro University, Indonesia

²Laboratorium Rekayasa Genetika, Pusat Antar Universitas Bioteknologi, Institut Teknologi Bandung, Indonesia

Published: 1 Apr 2005.

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Abstract

Polymerase Chain Reaction (PCR) merupakan metode amplifikasi Deoxyribonucleic acid (DNA) secara invitro. Hingga saat ini para peneliti baru berhasil mengamplifikasi secara in vitro DNA mitokondria (mtDNA)manusia Indonesia sampai ukuran fragmen maksimal 1,6 kilobasa (kb) dan belum dilakukan untuk “banyakfragmen” (multifragments). Pada penelitian ini telah berhasil diamplifikasi multifragments DNA mitokondriamanusia Indonesia berukuran 5,4 kb; 4,2 kb; 3,5 kb; 2,1 kb; 1,6 kb dengan metode PCR standar melalui optimasikonsentrasi MgCl2 dan suhu annealing. Proses berlangsung dalam satu reaksi PCR menggunakan enzim taqpolimerase, lima primer yaitu dmt-2L, dmt-1H, FL, FH dan MH pada kondisi bufer standar dengan konsentrasiMgCl2 2,5 mM, selama 30 siklus pada suhu denaturasi 94oC (15 detik), annealing 65oC (30 detik) danextension 72oC (180 detik). Keberhasilan metode amplifikasi in vitro multifragments yang mengandung fragmenpanjang tersebut diharapkan bisa membuat proses amplifikasi beberapa gen dalam satu genom menjadi lebihcepat dan efisien.

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Keywords: PCR multifragments; mtDNA manusia Indonesia; optimasi PCR; MgCl2

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Section: Research Articles

Language : ID

In Vol 8, No 1 (2005): Volume 8 Issue 1 Year 2005

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