In order to isolate Bacillus stearothermophillus , fifty soil samples werelected from the southern region of Iraq representing five Iraqi provinces luding Basrah , Thiqar , Misan , AL_Mothanna , and Karbalaa respectively .m these soil samples, fifty four local isolates were obtained and subjected to ntification according to their morphological, cultural, and biochemical tests.Results showed that twenty nine of these isolates were identified as cillus stearothermophillus. The ability of these local isolates in lipase duction was examined on LB-trybutyrin agar medium. Results showed that o of these isolates were lipase producers according to the diameters of drolysis zones. B.stearothermophillus BSR3 was selected for the molecular dy because it was efficient in the production of thermostable lipase.In attempt to clone thermostable lipase gene of BSR3 isolate in most miliar gram negative host (E. coli), lipase gene was amplified using specificprimers designed to include genetic elements upstream and downstream lipase gene located on chromosomal DNA of standard strain of B. stearothermophillus (U78785) according to the bioinformatics database available on NCBI web site .Results showed the amplified fragment was most identical (67% identity) to portion of nitrate reductase of Geobacillus sp.WCH70. As a consequence of this result new sets of primers were designed according to the complete sequence of lipase gene of Geobacillus sp.WCH70 to amplify lipase gene of BSR3 to consist of specific control sequences upstream and downstream of lipase gene. The amplified gene fragment was cloned in pPCRScriptSK (+) cloning vector and sequenced according to chain termination method. Results showed that the amplified lipase gene was identical (99% identity) to lipase gene of Geobacillus sp.WCH70. According to results of biochemical tests and DNA sequencing of lipase gene, locally isolated B. stearothermophillus BSR3 was regarded as Geobacillus stearothermophillus BSR3.
