UJI MUTU PRODUK BIOFORMULASI BAKTERI PENGENDALI SISTA Globodera rostochiensis DENGAN ANALISIS MOLEKULAR

AJENG DARA PRAMITA, Ir. Donny Widianto, Ph.D; Dr. Ir. Siwi Indarti, MP

2016 | Tesis | S2 Bioteknologi

Metode quantitave PCR dengan modifikasi propidium monozide telah dapat digunakan untuk membedakan jumlah sel hidup dan mati sehingga memungkinkan untuk dimanfaatkan dalam uji mutu produk bioformulasi. Tujuan dari penelitian ini adalah untuk mengetahui kemampuan teknik molekular dalam pengujian mutu produk bioformulasi agens hayati dan pengaruh bioformulasi bakteri terhadap efektifitas pengendalian sista G.rostochsiensis. Inokulum produk bioformulasi dipilih berdasarkan hasil karakterisasi, uji aktivitas enzim, dan uji perusakan sista oleh enzim kasar. Strain terpilih diformulasi pada bahan pembawa tepung beras, tepung pollard gandum dan tepung gaplek dan disimpan selama tiga bulan. Akurasi metode PMA-qPCR diuji dengan penentuan jumlah sel viabel pada sampel penyimpanan bulan ke 0 & 3. Efektivitas pengendalian bioformula strain-strain Bacillus terhadap sista NSK diuji secara in vitro dan in vivo pada skala mesokosmos. Perhitungan jumlah sel viabel menggunakan metode konvensional dan molekular menunjukkan jumlah sel pada kultur campuran lebih tinggi daripada kultur tunggal, yaitu sebesar log 10,54 sel/gram pada metode qPCR dan 7,38 pada metode TPC. Konsistensi hasil dari kedua metode uji membuktikan kemampuan metode qPCR dalam perhitungan jumlah sel viabel. Hasil uji perusakan sista oleh enzim kasar bioformula strain Bacillus menunjukkan kerusakan sista paling tinggi terdapat pada perlakuan kultur campuran yaitu sebesar 66%, diikuti oleh DN 42 sebesar 65% dan paling rendah pada DN 38 dan DN 44 yaitu sebesar 61,6%. Uji kontras terhadap tingkat kerusakan sista NSK tersebut menunjukkan kombinasi strain tidak memberikan pengaruh yang berbeda. Kesimpulan dari penelitian ini adalah teknik PMA-qPCR dapat digunakan dalam pengujian mutu produk bioformulasi agens hayati dan jumlah strain dalam produk agens hayati tidak memberikan pengaruh terhadap kemampuan pengendalian sista G.rostochiensis.

The ability of quantitative PCR technique with propidium Monoazide modification (PMA - qPCR ) to distinguish death and live cell allowing this method to be used in the quantification method of viable cell in bioformulation product. The objectives of this research are to determine the ability of molecular techniques to be used in quantification of viable cell in the bioformulation product and also to figure the influence of bacterial bioformulation in the efficacy of the G.rostochsiensis cyst's control ability. This research is conducted by doing a several experiment using biological and molecular assay. The determintion of viable cell in bacterial bioformulation product are performed in two methods that is conventional and molecular assay. To figuring the effects of bioformulation in the effication of biocontrol ability on golden cyst nematode, some tests are conducted using in vitro and in vivo bioassay. Total viable number of cell in mix culture formulation are higher than single culture formulation. These result are performed in both quantification method that is 10,54 log cells/gram in the qPCR method and 7,38 at TPC method. The consistency of these results in both quantification methods showing the ability of qPCR in quantifying viable cell. Result performed from PCN cyst destruction by Bacillus strain bioformula's crude enzyme showing the highest PCN cyst damage is performed by mixed cultures treatment, with the percentage of 66%, followed by DN 42 at 65% and the lowest result in DN 38 and DN 44 that is 61,6 %. Contrast test of PCN cyst damaged level from both single and mixed culture showing the combinations of strains did not gave any influence to the biocontrol efficacy. The conclusion of this study is the PMA - qPCR technique can be used in the quantification process bacterial bioformulation product. The amount of strain formulated in the biological agent's product did not give any influence on the ability of bacteria to control G.rostochiensis cyst.

Kata Kunci : Uji Mutu, Bioformulasi, sista, Globodhera rostochiensis, PMA, quantitative PCR.