ANALISIS URUTAN NUKLEOTIDA FRAGMEN GEN PENGKODE LIPASE DARI BAKTERI Alcaligenes sp. JG3

PENI LESTARINI, Dr. Tri Joko Raharjo, M.Si. ; Dr. Endang Astuti, M.Si.

2017 | Tesis | S2 Ilmu Kimia

Penelitian ini bertujuan untuk uji konfirmasi isolat bakteri, analisis urutan nukleotida fragmen gen pengkode lipase dan analisis struktur 3D lipase bakteri Alcaligenes sp. JG3. Konfirmasi isolat bakteri Alcaligenes sp. JG3 menggunakan gen 16S rRNA. Primer didesain dengan menggunakan software Primer Blast berdasarkan urutan gen pengkode lipase bakteri Alcaligenes faecalis subsp. faecalis NCIB 8687. Sekuensing hasil amplifikasi DNA secara PCR dilakukan dengan genetic analyzer. Urutan nukleotida gen pengkode lipase bakteri Alcaligenes sp. JG3 disejajarkan untuk mengetahui persentase kemiripan dengan urutan nukleotida acuan. Prediksi struktur 3D menggunakan server I-TASSER. Analisis menggunakan gen 16S rRNA mengkonfirmasi bahwa isolat bakteri memiliki persentase kemiripan 99% dengan bakteri spesies Alcaligenes sp. Pasangan primer foward 5'-CATGAGCCTGAACAAAGGTG-3' dan reverse 5'- TACATGCTGGTGGATAGCTG-3' mampu mengamplifikasi DNA gen pengkode lipase bakteri Alcaligenes sp. JG3 menghasilkan ukuran fragmen 864 pb (0,8 kb). Fragmen 0,8 kb gen pengkode lipase bakteri Alcaligenes sp. JG3 memiliki persentase kemiripan sebesar 91% dan 87,5% terhadap urutan nukleotida lipase Alcaligenes faecalis strain ZD 02 dan Alcaligenes faecalis subsp. faecalis NCIB 8687. Fragmen tersebut menunjukkan kemiripan identitas struktur 3D dan sisi aktif terhadap Alcaligenes faecalis subsp. faecalis NCIB 8687.

This research aimed to confirm the bacterial isolate, to perform nucleotide sequencing encoding lipase and to analysis the 3D structure of lipase gene from Alcaligenes sp. JG3 bacterium. The isolate confirmation of Alcaligenes sp. JG3 has been done using 16S rRNA gene. Primers were designed based on the nucleotide sequence encoding lipase present in Alcaligenes faecalis subsp. faecalis NCIB 8687. The DNA sequencing was performed with a genetic analyzer. Nucleotide sequence encoding lipase of Alcaligenes sp. JG3 was aligned to know its percentage of similarity with reference nucleotide sequence. The model of 3D structure was predicted using I-TASSER. The analysis of 16S rRNA gene confirmed that the bacterial isolates had 99% similarity towards Alcaligenes sp. The selected primers were foward 5'-CATGAGCCTGAACAAAGGTG-3' and reverse 5'-TACATGCTGGTGGATAGCTG-3 could amplify the DNA template of this bacterium resulting 864 pb (0.8 kb) respectively. The sequence analysis showed that the fragment had 91% similarity towards Alcaligenes faecalis strain ZD 02. The 3D structure and active side showed that the fragment had similarity towards Alcaligenes faecalis subsp. faecalis NCIB 8687.

Kata Kunci : Alcaligenes sp. JG3, gen lipase, I-TASSER