苏云金芽孢杆菌肠毒素基因entFM的定位与序列分析

Localization and Characterization of entFM Gene of Bacillus thuringiensis

为明确BtWB9肠毒素基因entFM在基因组中的位置及该基因的特性,以entFM基因片段制备探针,通过Southern杂交,分别对染色体DNA和质粒DNA进行基因杂交定位;以PCR方法克隆了BtWB9、1072、TS16及Bc6A1entFM基因ORF序列,并运用生物信息学工具进行了核苷酸序列与推导氨基酸序列分析。基因定位结果表明,BtWB9的entFM基因位于染色体上;基因序列分析表明,WB9entFM基因的ORF序列共由1281个核苷酸组成,可编码426个氨基酸。由基因序列推导的氨基酸序列分析显示,WB9、TS16、1072与BcCx5之间主要差异在于BcCx5的EntFM氨基酸序列在39～42位置多出QTQT(谷氨酰胺-苏氨酸-谷氨酰胺-苏氨酸)4个氨基酸,在96～99位置还有SWDK4个氨基酸的差异,而其相似性都在92%以上。

The objectives of the study were to determine the location of entFM gene of WB9 in genome and the characterization of entFM gene sequence. The probe for entFM gene of WB9 was prepared and hybridized with plasmid DNA and chromosomal DNA of WB9 respectively. The ORF of entFM genes in Bt WB9, TS16, 1072 and Bc 6A1 were cloned, sequenced and analyzed by tools of bioinformatics. The results indicated that entFM gene of WB9 was in the chromosome. Sequence analysis showed that entFM gene of WB9 contained 1281 nucleotide acids, encoding 426 amino acids. The comparison of the predicted amino acid sequences showed that the main differences between WB9 and Cx5 were that 4 amino acids （Gln-Thr-Gln-Thr） were deleted between 39th and 42th in WB9 and 4 different amino acids （SWDK） were added between 96th and 99th in Be Cx5. The amino acid sequences identities were over 92% between WB9, TS16, 1072 and Cx5.