BAKTERI ENDOFIT TANAMAN PURWOCENG (Pimpinella pruatjan Molk.) BERDASARKAN KARAKTER MORFOLOGIS, BIOKIMIAWI DAN MOLEKULAR

WILDIANI WILSON, Dr. Yekti Asih Purwestri, S.Si, M.Si

2014 | Tesis | S2 Biologi

Isolasi, karakterisasi, identifikasi dan screening antimikrobia bakteri endofit dari akar tanaman purwoceng (Pimpinella pruatjan Molk.) telah dilakukan. Isolasi bakteri endofit dilakukan dengan metode irisan segmen akar. Karakterisasi bakteri endofit dilakukan berdasarkan morfologis, biokimiawi dan molekular. Karakter morfologis dapat dilihat melalui bentuk koloni, bentuk sel, tepian, elevasi dan warna koloni. Karakter biokimiawi dilakukan dengan uji fermentasi karbohidrat, katalase, hidrolisis pati, produksi H2S, reduksi nitrat dan uji oksidase. Karakter molekular didasarkan pada gen 16S rRNA. DNA genom bakteri yang didapatkan diamplifikasi dengan PCR menggunakan primer spesifik untuk gen 16S rRNA lalu dikarakterisasi lebih lanjut berdasarkan metode amplified rDNA restriction analysis (ARDRA) menggunakan enzim restriksi AluI, RsaI, HaeIII dan HinfI. Pengujian aktivitas antimikrobia bakteri endofit menggunakan metode blok agar. Sebanyak 69 koloni bakteri endofit berhasil diisolasi dan hanya 30 isolat berhasil dipurifikasi. Isolat yang diperoleh didominasi oleh bakteri Gram negatif (70%). Hasil karakter morfologis dan biokimiawi menunjukkan keanekaragaman isolat bakteri endofit. Tiga puluh isolat bakteri endofit diidentifikasi sebagai genus Acidomonas, Amphibacillus, Arsenophonus, Bacillus, Brocothrix, Budvicia, Cardiobacterium, Cellulomonas, Erwinia, Pantoea, Pediococcus, Plesiomonas dan Proteus. Screening aktivitas antimikrobia menunjukkan hanya isolat GP11 dan GP12 yang berpotensi menghambat Staphylococcus aureus dengan zona hambat yang sama yaitu 1,72 cm. Hasil fingerprinting DNA melalui pemotongan oleh enzim RsaI dan HaeIII mampu menunjukkan isolat potensial antimikrobia merupakan strain yang berbeda. Penelitian ini menunjukkan bahwa bakteri endofit dapat ditemukan pada akar purwoceng dan memiliki potensi antimikrobia.

Isolation, characterization, identification and antimicrobial screening of endophytic bacteria from purwoceng’s roots (Pimpinella pruatjan Molk.) has been carried out. Isolation of endophytic bacteria was conducted using slices of the root segments. The characterization of endophytic bacteria was conducted based on morphological, biochemical and molecular characters. Morphological characters can be seen through the form of colonies, cell shape, edges, elevation and color of the colony. Biochemical characters were performed by carbohydrate fermentation, catalase, starch hydrolysis, H2S production, nitrate reduction and oxidase test. Molecular character based on 16S rRNA gene. Bacterial genomic DNA obtained was amplified by PCR using specific primers for the 16S rRNA gene and further characterized by amplified rDNA restriction analysis method (ARDRA) using restriction enzymes AluI, RsaI, HaeIII and HinfI. Antimicrobial activity of endophytic bacteria using agar block method. A total of 69 colonies of endophytic bacteria were isolated and only 30 isolates were successfully purified. Isolates obtained were dominated by Gram-negative bacteria (70%). The results of morphological and biochemical characters showed the diversity of endophytic bacteria isolates. Thrity isolates of endophytic bacteria were identified as Acidomonas, Amphibacillus, Arsenophonus, Bacillus, Brocothrix, Budvicia, Cardiobacterium, Cellulomonas, Erwinia, Pantoea, Pediococcus, Plesiomonas and Proteus. Screening for antimicrobial activity showed only isolate GP11 and GP12 were potentially inhibit Staphylococcus aureus with the same inhibition zone is 1,72 cm. Result of fingerprinting DNA by enzymes RsaI digestion products and HaeIII were able to demonstrate the potential isolates are different strains. This study suggests that endophytic bacteria can be found in the roots and have the potential antimicrobial

Kata Kunci : P. pruatjan Molk., bakteri endofit, Antimikrobia, ARDRA