Identifikasi Molekuler dan Desain Primer Spesifik Gen rpoB pada Bakteri Streptomyces sp. strain UGM_P-I2

Diah Ayu Wulandari, Prof. Dr. Ir. Siti Subandiyah, M.Agr.Sc.; Dr. Ir. Arif Wibowo, M.Agr.Sc.

2026 | Skripsi | ILMU HAMA & PENYAKIT TUMBUHAN

Genus Streptomyces mencakup hampir sebagian besar bakteri Actinomycetes yang dikenal sebagai penghasil metabolit sekunder dengan peran beragam sebagai antibakteri, antijamur, insektisida, herbisida, dan perangsang pertumbuhan tanaman. Produksi metabolit sekunder dikendalikan oleh enzim – enzim yang dikodekan biosynthetic gene clusters (BGCs) yang dapat spesifik pada tingkat spesies hingga strain. Streptomyces sp. strain UGM_P I2 diketahui menghasilkan senyawa antifungi dan insektisida. Kemampuan dalam menghasilkan senyawa tersebut berkaitan dengan regulasi fisiologisnya yang dipengaruhi oleh gen housekeeping seperti gen rpoB yang bersifat konservatif. Penelitian ini bertujuan merancang primer spesifik untuk mendeteksi gen rpoB pada isolat Streptomyces sp. strain UGM_P-I2 dan mengidentifikasi filogeni isolat tersebut. Desain primer spesifik gen rpoB dilakukan secara in silico menggunakan Geneious Primer, kemudian diverifikasi dengan Polymerase Chain Reaction (PCR), dan analisis filogenetik dilakukan menggunakan MEGA 12. Primer spesifik yang telah dirancang dapat mendeteksi gen rpoB pada isolat Streptomyces sp. strain UGM_P-I2 dengan amplikon berukuran 221 bp pada suhu annealing 63,2°C. Primer spesifik yang berhasil dirancang adalah Strep-rpoBF (5’-CTGAACGAGGAGATGCGGT-3’) dan Strep-rpoBR (5’-GACTTGATCAGCGGTACG-3’). Analisis filogenetik menunjukkan bahwa Streptomyces sp. strain UGM_P-I2 memiliki kekerabatan dekat dengan Streptomyces broussonetiae strain T44.

The genus Streptomyces comprises most Actinomycete bacteria and is well known as a producer of secondary metabolites with diverse roles as antibacterial, antifungal agents, insecticides, herbicides, and plant growth promoters. The production of these secondary metabolites is controlled by enzymes encoded by biosynthetic gene clusters (BGCs), which can be specific at the species and even strain level. Streptomyces sp. strain UGM_P I2 is known to produce antifungal and insecticidal compounds. The ability to produce these compounds is related to their physiological regulation, which is influenced by housekeeping genes such as the conserved rpoB gene. This study aimed to design specific primers to detect the rpoB gene in Streptomyces sp. strain UGM_P I2 and identify the phylogeny of this isolate. Specific primers for the rpoB gene were designed in silico using Geneious Primer, then verified by Polymerase Chain Reaction (PCR), and phylogenetic analysis was performed using MEGA 12. The designed specific primers were able to detect the rpoB gene in Streptomyces sp. strain UGM_P I2 with an amplicon size of 221 bp at an annealing temperature of 63.2 °C. The specific primers successfully designed were Strep rpoBF (5'-CTGAACGAGGAGATGCGGT-3') and Strep rpoBR (5'-GACTTGATCAGCGGTACG-3'). Phylogenetic analysis showed that Streptomyces sp. strain UGM_P I2 is closely related to Streptomyces broussonetiae strain T44.

Kata Kunci : Streptomyces sp., gen rpoB, desain primer, PCR, in silico