类似S-层蛋白的苏云金芽胞杆菌伴胞晶体蛋白基因的克隆

芽胞杆菌CTC菌被鉴定为苏云金芽胞杆菌，鞭毛血清型H2，幕虫亚种；产生卵圆形伴胞晶体，伴胞晶体蛋白为100kD；测定了该蛋白 N－末端序列，该序列与炭疽芽胞杆菌的细胞表面S－层蛋白具92－93％相似性，根据Southern杂交制作了该晶体蛋白基因ctc所在位置的限制性酶切图谱，分别克隆了该基因5’和3’端所在2．9kb XbaI片段和3．1kb Cla I DNA片段，彼此间具0．6kb重叠，通过拼接获得含完整ctc基因的克隆，含该基因的大肠杆菌与表达S－层蛋白的大肠杆菌具相似生长特征，初步表明CTC菌株的伴胞晶体由细胞表面S－层蛋白组成，苏云金芽胞杆菌区别于蜡状芽胞杆菌和炭疽芽胞杆菌的唯一标准是能形成伴胞晶体，由于S－层是细胞表面的结构成分，本文对CTC菌株鉴定为苏云金芽胞杆菌以及伴胞晶体作为苏云金芽胞杆菌鉴别的唯一标准提出了质疑。

CLONING OF PARASPORAL BODY PROTEIN GENE RESEMBLING TO S-LAYER PROTEIN GENES FROM BACILLU THURINGIENSIS CTC STRAIN

Bacillus CTC strain was identified as Bacillus thuringiensis subsp. finitimus (serotype H2) and Pasteur Institute confirmed this identification. The parasporal body formed by CTC strain is oval shaped, and consists of 100 kD protein. The determination of the N-terminal amino acid sequence showed this protein shares 93% similarity to that of B. anthracis S-layer proteins. The restriction map covering the related protein gene (ctc) was deduced according to southern hybridization. The DNA fragments containing the 5' and 3' end of ctc gene with 0.6 kb overlap were isolated, respectively. Full length ctc gene was then constructed. B. thuringiensis is a species similar to B. cereus and B. anthracis, and only can be distinguished by forming parasporal bodies. The growth of E. coli maintaining ctc gene is similar to that of E. coli expressing S-layer protein gene. The above preliminary results showed that the parasporal body protein in CTC strain is a cell surface-like protein. So identifying CTC strain as a species of B. thuringiensis and the unique criterion for the identification of B. thuringiensis by forming parasporal body should be reconsidered.