VOLUME 5 NUMBER 2 (July to December 2012)

Philipp. Sci. Lett. 2012 5 (2) 209-215
available online: December 19, 2012

*Corresponding author
Email Address: wlrivera@science.upd.edu.ph
Submitted: August 24, 2012
Revised: November 05, 2012
Accepted: November 05, 2012

ARTICLE

Isolation of thermophilic bacteria(Bacillus AND Ureibacillus)and amplification of genes forselected enzymes

by Cynthia T. Hedreyda*, John Jewish A. Dominguez, and Karen Rosal

National Institute of Molecular Biology and Biotechnology, College of Science,
     University of thePhilippines, Diliman, Quezon City, Philippines 1101
Because catalyses often employ or produce hightemperature environment, isolation of bacterialthermophiles that produce thermostable enzymesis important in industry. This study was focusedon the isolation of bacteria that can grow at 55°Cand higher, by using hot spring, mud spring, and solid oil sludgesamples as sources of inocula added to Luria Bertini broth thatwere incubated at 55 to 70°C with shaking (25 rpm). One isolatefrom the hot spring sample that grew at 60°C and another isolatefrom oil sludge sample that grew at 55°C, were identified to belong to Bacillus licheniformis. An isolate from mud spring thatgrew at 55°C was identified to belong to Bacillus subtilis. Pureculture of two isolates from the oil sludge sample that couldsurvive up to 60°C exhibited 99% 16s rRNA gene sequencesimilarity with two relatively new species, Ureibacillussuwonensis and Ureibacillus thermosphaericus. The apr foralkaline protease was detected from the mud spring B.licheniformis while a fragment of a gene with 98% sequencesimilarity with the gene for Bacillopetidase F was amplifiedfrom the B. licheniformis isolated from oil sludge. PCR usinggene-targeted primers and template DNA of the Bacillus subtilisisolate from mudspring, resulted in the amplification of expectedsize amplicons for a neutral protease, β-glycosidase, α-amylaseand xylanase genes. Amplification of target gene fragments wasconfirmed by gene sequence analysis. Target amplicons were notgenerated from the Ureibacillus suwonensis and Ureibacillusthermosphaericus isolates, suggesting the need for furtherstudies using new or degenerate primers to amplify genes forenzymes with industrial applications. These relatively two newbacterial species that are not yet well studied, particularly at themolecular level, could be potential sources of important enzymesthat allow the bacteria to survive and grow in extremeenvironments such as the oil sludge.

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