GENETIC DI VERSITY AN ALYSIS OF THERMOPHILIC BACTERIA FROM C ANDRADIMUKA CR ATER IN CENT RAL J AVA EMPLOYING PCR-RFLP OF 16S- rRNA GEN E

Abstract

The specific primers for bacteria (63f a nd 1387r) were use d to amplify the 16 S-rRNA genes fro m total com munit y genom ic DNA of t herm ophi lic bacteri a. The t otal communit y genomic DNA was obtained from mud s and wate r sa mples of Candrad imuka crater, Dieng P lateau, Centra l Ja va. PCR products were cloned into vector pCR*2.1-TOPO (3.9 kb) and transformed into Escherichia coli TOPIC. Tw o tetrameric restriction endonucleases Rsal and Hhal were employed to generate Restric tion Fra gment Length Polymorphisms (RFLP) p aterns . T hese enzym es yielded 10 and 9 groups of 16S-rRNA pro files or OTU (Opera tion al Taxonomic Un its ) from 27 1 6S-rRNA ge ne c lones. Rsal w as fo un d to be mo re d iscriminativ e in diff er entiat in g the cl on es t han Hhal. R sal-RFLP indi cated that OTU 7 and OTU 3 represented the most abundant clones, i.e. 6 and 5 clones respective ly. The distribution of 16S-r RNA gene c lones could indicate relati ve dist ri butio n of specifi c groups of thermophili c bacteria in th ei r natural h abitat. Analysis o f diversity at the DNA level could represent both culturable and unculturabl e bacteria in t he envir onment . Similarity analysis sho wed that at level 0.600 there were 8 diff erent groups from 1 0 RFLP profiles genera te d by Rsal digestion. This study in dicated tha t the re were at leas t 8 grou ps of different thermoph ilic bacter ia occupying Candradimuka crater.

Keywords

• Thermophile s
• 16S-rRNA
• Candradimu ka crater.