Andrup L, Damgaard J, Wassermann K, Boe L, Madsen S M, Hansen F G
National Institute of Occupational Health, Department of Toxicology and Biology, Copenhagen, Denmark.
Plasmid. 1994 Jan;31(1):72-88. doi: 10.1006/plas.1994.1008.
The complete nucleotide sequence of the plasmid pTX14-3 from Bacillus thuringiensis subsp. israelensis has been determined. The circular DNA molecule was 7649 bp and had a G + C content of 35.1%. Twenty-two open reading frames larger than 50 codons were identified. Ten of these open reading frames are suggested to be protein coding regions. The existence of the polypeptides encoded by the mob14-3 and rep14-3 genes were verified by maxi-cells analysis in Escherichia coli. Even though the rep14-3 gene was expressed in E. coli the plasmid pTX14-3 was unable to replicate in this bacterium. The minimal region of the plasmid pTX14-3 required for replication in B. thuringiensis was identified. Potential secondary structures upstream of the rep14-3 gene indicated regulation by antisense RNA and transcription attenuation. Extensive sequence homology with the B. thuringiensis subsp. thuringiensis plasmid pGI2 was found in the last part of the mob14-3 gene, downstream of the rep14-3 gene, and in the region containing the single-strand origin of replication (i.e., the minus origin) of pTX14-3. A sequence of 700 bp containing multiple direct repeats was found in an ORF encoding a glycine and proline rich protein of 35.9 kDa. 1.2 kbp upstream and 0.1 kbp downstream of this ORF was found a large direct repeat of 230 bp (87% identity). The region between this direct repeat was often spontaneously deleted from plasmid derivatives containing the entire pTX14-3.
苏云金芽孢杆菌以色列亚种质粒pTX14 - 3的完整核苷酸序列已被测定。该环状DNA分子为7649 bp，G + C含量为35.1%。鉴定出22个大于50个密码子的开放阅读框。其中10个开放阅读框被认为是蛋白质编码区。通过在大肠杆菌中的大细胞分析验证了mob14 - 3和rep14 - 3基因编码的多肽的存在。尽管rep14 - 3基因在大肠杆菌中表达，但质粒pTX14 - 3在该细菌中无法复制。确定了质粒pTX14 - 3在苏云金芽孢杆菌中复制所需的最小区域。rep14 - 3基因上游的潜在二级结构表明存在反义RNA调控和转录衰减。在mob14 - 3基因的最后部分、rep14 - 3基因的下游以及包含pTX14 - 3单链复制起点（即负链起点）的区域，发现与苏云金芽孢杆菌苏云金亚种质粒pGI2有广泛的序列同源性。在一个编码35.9 kDa富含甘氨酸和脯氨酸蛋白的开放阅读框中发现了一个包含多个直接重复序列的700 bp序列。在该开放阅读框上游1.2 kbp和下游0.1 kbp处发现了一个230 bp的大直接重复序列（87%同源性）。在含有完整pTX14 - 3的质粒衍生物中，这个直接重复序列之间的区域经常自发缺失。
