SILENCING mac-miR156m MENARGETKAN GEN di Fusarium oxysporum f. sp. cubense Tropical Race 4 (Foc TR4) SEBAGAI PENGHAMBAT PERTUMBUHAN JAMUR PENYEBAB LAYU PISANG CAVENDISH

Rina Budi Astuti, Prof. Dr. Ir. Siti Subandiyah, M.Agr.Sc ; Dr. Ir. Arif Wibowo, M.Agr.Sc

2024 | Tesis | S2 Bioteknologi

Fusarium oxysporum f.sp cubense Tropical Race 4 (Foc TR4) merupakan patogen virulen penyebab hilangnya hasil pisang Cavendish akibat infeksi di seluruh area tanam. Beberapa tahun terakhir, mekanisme interferensi RNA (RNAi) menggunakan microRNAs (miRNAs) menjadi fokus penelitian mengenai interaksi antara tanaman dan patogen, serta pengembangan alternatif ramah lingkungan dalam pengendalian patogen. Penelitian ini, menggunakan analisis ekspresi gen dan pendekatan analisis in silico pencarian kandidat miRNA (mac-miR156m) menggunakan website seperti miRBase, TarDB, dan psRNAtarget. mac-miR156m yang disintesis kemudian diaplikasikan kedalam koloni jamur dengan penetesan menggunakan mikropippet, dilihat morfologi serta analisis ekspresi gen target pada jamur Foc TR4. Hasil pemberian mac-miR156m terlihat lebih menekan pada sporulasi jamur sebesar 66,8% pada konsentrasi 20 ng/µl dan 75,7% pada konsentrasi 20 ng/µl dibandingkan diameter koloni dengan penghambatan sebesar 4,45?n 4,43% pada konsentrasi 20 dan 50 ng/µl selama 20 hari pengamatan. Penurunan ekspresi relatif gen target pada konsentrasi 20 ng/µl ARB 95 % (19,56 kali), LETM 94% (16,06 kali), PM 89% (8,78 kali) dan SIX1C 62% (2,67 kali) menurunkan ekspresi gen lebih baik dibandingkan konsentrasi 50 ng/µl. Pola ekspresi miRNA yang ditentukan dengan metode kuantifikasi menunjukkan bahwa miRNA memainkan peran penting dalam patogenisitas Foc TR4.

Fungal pathogens are the largest group of agents that attack plants. Fusarium oxysporum f.sp cubense Tropical Race 4 (Foc TR4) is a virulent pathogen that caused Cavendish banana yield loss due to infection throughout the planting area. In recent years, RNA interference (RNAi) mechanisms have been the main focus of research on the interactions between plants and pathogens, and the development of an environmental alternative to pathogen control. small RNA is involved in silencing genes known to be transmitted bidirectionally between fungal pathogens and their hosts. microRNAs (miRNAs) have been identify in several plants and play important roles. However, the application of miRNAs from plants directly to the pathogen Foc TR4 has not been reported. In this study, we used the gene expression analysis and bioinformatics analysis approach. The total miRNA family of plants selected by in silico analysis as miRNA candidate, that is MiR156 from miRBase and selected mac-miR156m as candidate sequences for gene inhibition test in Foc TR4. RT-qPCR analysis was used for validation of miRNA-targeted genes, a total of 4 genes were confirmed their expression. The miRNA expression patterns determined by the quantification method suggested that miRNAs play an important role in the pathogenicity of extremely virulent Foc TR4.

Kata Kunci : Interference RNA, microRNA, mac-miR156m, Foc TR4