Pemanfaatan Teknologi Droplet Digital PCR (ddPCR) dalam Kegiatan Analisis Molekuler Tanaman

Abstract

(Article History: Received 23 October 2020; Revised 9 January 2021; Accepted 18 January 2021)

ABSTRAK

Selama beberapa dekade terakhir, teknik PCR memberikan manfaat yang begitu besar dalam kegiatan penelitian di bidang biologi molekuler. Digital droplet PCR (ddPCR) merupakan salah satu teknologi PCR terbaru yang diklaim memiliki keunggulan dibanding teknik qPCR. Prinsip kerja teknik ini yaitu membagi sampel menjadi molekul-molekul kecil yang dipisahkan oleh emulsi minyak, air, dan senyawa penstabil sehingga membentuk droplets. Teknik ini memiliki kelebihan mampu melakukan kuantifikasi absolut maupun relatif pada DNA dengan konsentrasi sangat rendah, tidak memerlukan kurva standar, serta tidak sensitif terhadap kehadiran senyawa inhibitor. Teknik ini telah diaplikasikan pada kegiatan analisis molekuler tanaman di antaranya kegiatan pengukuran konsentrasi DNA dengan sangat akurat, deteksi kehadiran patogen pada jaringan tanaman, dan estimasi jumlah salinan T-DNA pada proses transformasi genetik.

Kata kunci: PCR; droplet digital PCR; DNA; biologi molekuler; alat deteksi

ABSTRACT

Over the past decades, PCR technique has provided enormous benefits in molecular biology research activities. Digital droplet PCR (ddPCR) is one of the latest PCR technologies that is claimed to have advantages over the qPCR technique. The working principle of this technique is to divide the sample into small molecules, which separated by emulsions of oil, water, and stabilizing compounds to form droplets. This technique has the advantage of being able to perform absolute and relative quantification with very low DNA concentrations, does not require a standard curve, and less sensitive to the presence of inhibitor compounds. This technique has been applied to a number of plant molecular analysis, such as for measuring DNA concentrations very accurately, detecting the presence of pathogens in plant tissue, and estimating the copy number of T-DNA in the genetic transformation process.

Keywords: PCR; droplet digital PCR; DNA; molecular biology; diagnostic tool.