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Li, Ming Shun, Huazhong Agricultural University, Wuhan, P. R. China; Roh, J.Y., Seoul National University, Seoul, Republic of Korea; Tao, Xueying, Seoul National University, Seoul, Republic of Korea; Yu, Zi Niu, Huazhong Agricultural University, Wuhan, P. R. China; Liu, Zi Duo, Huazhong Agricultural University, Wuhan, P. R. China; Liu, Qin, Huazhong Agricultural University, Wuhan, P. R. China; Xu, Hong Guang, Huazhong Agricultural University, Wuhan, P. R. China; Shim, H.J., Seoul National University, Seoul, Republic of Korea; Kim, Y.S., Seoul National University, Seoul, Republic of Korea; Wang, Y., Seoul National University, Seoul, Republic of Korea; Choi, J.Y., Seoul National University, Seoul, Republic of Korea; Je, Y.H., Seoul National University, Seoul, Republic of Korea
The journal of microbiology, 08/2009, Letnik: 47, Številka: 4Journal Article
Bacillus thuringiensis, an entomopathogenic bacterium belonging to the B. cereus group harbors numerous extra-chromosomal DNA molecules whose sizes range from 2 to 250 kb. In this study, we used a plasmid capture system (PCS) to clone three small plasmids from R thuringiensis subsp. kurstaki K1 which were not found in B. thuringiensis subsp. kurstaki HD-1, and determined the complete nucleotide sequence of plasmid pK1S-1 (5.5 kb). Of the six putative open reading frames (ORF2-ORF7) in pK1S-1, ORF2 (MobK1) showed approximately 90% aa identity with the Mob-proteins of pGI2 and pTX14-2, which are rolling circle replicating group Ⅶ (RCR group Ⅶ) plasmids from B. thuringiensis. In addition, a putative origin of transfer (oriT) showed 95.8% identity with those of pGI2 and pTX14-2. ORF3 (RepK1) showed relatively low aa identity (17.8 similar to 25.2%) with the Rep protein coded by RCR plasmids, however. The putative double-strand origin of replication (dso) and single-strand origin of replication (sso) of pK1S-1 exhibited approximately 70% and 64% identities with those of pGI2 and pTX14-2. ORF6 and 7 showed greater than 50% similarities with alkaline serine protease, which belongs to the subtilase family. The other 2 ORFs were identified as hypothetical proteins. To determine the replicon of pK1S-1, seven subclones were contructed in the B. thuringiensis ori-negative pHT1K vector and were electroporated into a plasmid cured B. thuringiensis strain. The 1.6 kb region that included the putative ORF3 (Rep1K), dso and ORF4, exhibited replication ability. These findings identified pK1S-1 as a new RCR group Ⅶ plasmid, and determined its replication region.
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