Identifikasi spesies isolat bakteri termofilik dari Kalianda Lampung Selatan dengan analisis urutan Gen 16s rRNA

Legiastuti, Tyas Mega (2022) Identifikasi spesies isolat bakteri termofilik dari Kalianda Lampung Selatan dengan analisis urutan Gen 16s rRNA. Sarjana thesis, UIN Sunan Gunung Djati Bandung.

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Abstract

Bakteri termofilik merupakan bakteri yang beradaptasi dengan kondisi lingkungan yang bersuhu tinggi, yaitu suhu berkisar 45-90C. Bakteri termofilik menghasilkan enzim yang bersifat termostabil sangat dibutuhkan di berbagai bidang industri. Penelitian ini bertujuan untuk mengidentifikasi spesies bakteri termofilik dari air panas way belerang Lampung Selatan secara fenotip dan genotip. Analisis secara fenotip pada tahap pertama dilakukan peremajaan untuk mendapatkan isolat murni, kemudian dilakukan pengamatan morfologi isolat dan sel dengan pewarnaan gram serta uji biokimia untuk menentukan genus dari isolat bakteri. Selanjutnya dilakukan analisis secara genotip dengan sekuensing fragmen gen 16s rRNA untuk mengidentifikasi spesies dari sampel bakteri termofilik. DNA kromosom dari bakteri Bacillus diisolasi dan diamplifikasi menggunakan instrumen Polymerase Chain Reaction (PCR) dengan primer forward 27 dan reverse 1492 untuk memperoleh fragmen gen 16s rRNA. Fragmen 16s rRNA yang diperoleh selanjutnya disekuensing untuk mengetahui urutan basa nukleotida metode dideoksi Sanger. Urutan nukleotida hasil sekuensing dimasukkan ke dalam program BLAST untuk mengetahui homologinya dengan urutan nukleotida fragmen 16s rRNA bakteri lain yang telah terdaftar di Bank Gen. Hasil pewarnaan gram menunjukkan isolat bakteri termofilik merupakan bakteri gram positif dan dari hasil uji biokimia didapatkan genus Bacillus. Hasil amplifikasi DNA kromosom isolat bakteri termofilik air panas way belerang kode Tyas_1b2 menunjukkan pita ± 1.500 bp. Hasil analisis urutan basa 16s rRNA menunjukkan isolat termofilik kode Tyas_1b2 memiliki indeks kemiripan sebesar 99,83% dengan Bacillus cereus. Thermophilic bacteria are bacteria that adapt to high-temperature environmental conditions, namely temperatures ranging from 45-90C. Thermophilic bacteria produce thermostable enzymes which are needed in various industrial fields. This study aims to identify the species of thermophilic bacteria from the Sulfur Hot Springs of South Lampung by phenotype and genotype. Phenotypic analysis was carried out in the first stage of rejuvenation to obtain pure isolates, then observed the morphology of isolates and cells with gram staining and biochemical tests to determine the genus of bacterial isolates. Furthermore, genotypic analysis was carried out by sequencing the 16s rRNA gene fragment to identify species from thermophilic bacteria samples. Chromosomal DNA from Bacillus bacteria was isolated and amplified using a Polymerase Chain Reaction (PCR) instrument with forward 27 and reverse 1492 primers to obtain 16s rRNA gene fragments. The 16s rRNA fragment obtained was then sequenced to determine the sequence of nucleotide bases using the dideoxy Sanger method. The sequenced nucleotide sequences were entered into the BLAST program to determine their homology with the nucleotide sequences of the 16s rRNA fragments of other bacteria that have been registered in the Gene Bank. The results of gram staining showed that the isolates of thermophilic bacteria were gram-positive bacteria and from the results of biochemical tests, the genus Bacillus. The results of amplification of chromosomal DNA isolates of thermophilic bacteria with sulfur code Tyas_1b2 showed a band of ± 1,500 bp. The results of the base sequence analysis of 16s rRNA showed that the thermophilic isolate with code Tyas_1b2 had a similarity index of 99,83% with Bacillus cereus.

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