Trisrisook M, Pantuwatana S, Bhumiratana A, Panbangred W
Department of Microbiology, Mahidol University, Bangkok, Thailand.
Appl Environ Microbiol. 1990 Jun;56(6):1710-6. doi: 10.1128/aem.56.6.1710-1716.1990.
A 3.7-kilobase (kb) XbaI fragment harboring the cryIVB gene (L. Thorne, F. Garduno, T. Thompson, D. Decker, M. A. Zounes, M. Wild, A. M. Walfield, and T. J. Pollock, J. Bacteriol. 166:801-811, 1986) which encoded a 130-kilodalton (kDa) mosquitocidal toxin from a 110-kb plasmid of Bacillus thuringiensis subsp. israelensis 4Q2-72 was cloned into pUC12 and transformed into Escherichia coli. The clone with a recombinant plasmid (designated pBT8) was toxic to Aedes aegypti larvae. The fragment (3.7 kb) was ligated into pBC16 (tetracycline resistant [Tcr]) and transformed by the method of protoplast transformation into Bacillus sphaericus 1593 and 2362, which were highly toxic to Anopheles and Culex mosquito larvae but less toxic to Aedes larvae. After cell regeneration on regeneration medium, the Tcr plasmids from transformants (pBTC1) of both strains of B. sphaericus were prepared and analyzed. The 3.7-kb XbaI fragment from the B. thuringiensis subsp. israelensis plasmid was shown to be present by agarose gel electrophoresis and Southern blot hybridization. In addition, B. sphaericus transformants produced a 130-kDa mosquitocidal toxin which was detected by Western (immuno-) blot analysis with antibody prepared against B. thuringiensis subsp. israelensis 130-kDa mosquitocidal toxin. The 50% lethal concentrations of the transformants of strains 1593 and 2362 against A. aegypti larvae were 2.7 X 10(2) and 5.7 X 10(2) cells per ml, respectively. This level of toxicity was comparable to the 50% lethal concentration of B. thuringiensis subsp. israelensis but much higher than that of B. sphaericus 1593 and 2362 (4.7 X 10(4) cells per ml) against A. aegypti larvae.(ABSTRACT TRUNCATED AT 250 WORDS)
一个含有cryIVB基因的3.7千碱基（kb）XbaI片段（L. 索恩、F. 加尔杜诺、T. 汤普森、D. 德克尔、M. A. 祖内斯、M. 怀尔德、A. M. 沃尔菲尔德和T. J. 波洛克，《细菌学杂志》166:801 - 811，1986年），该基因编码一种来自苏云金芽孢杆菌以色列亚种4Q2 - 72的110 kb质粒的130千道尔顿（kDa）杀蚊毒素，被克隆到pUC12中并转化到大肠杆菌中。带有重组质粒（命名为pBT8）的克隆对埃及伊蚊幼虫有毒性。该片段（3.7 kb）被连接到pBC16（四环素抗性[Tcr]）中，并通过原生质体转化方法转化到球形芽孢杆菌1593和2362中，这两种菌株对按蚊和库蚊幼虫具有高毒性，但对伊蚊幼虫毒性较小。在再生培养基上细胞再生后，制备并分析了两种球形芽孢杆菌菌株转化体（pBTC1）的Tcr质粒。通过琼脂糖凝胶电泳和Southern印迹杂交表明，来自苏云金芽孢杆菌以色列亚种质粒的3.7 kb XbaI片段存在。此外，球形芽孢杆菌转化体产生了一种130 kDa杀蚊毒素，用针对苏云金芽孢杆菌以色列亚种130 kDa杀蚊毒素制备的抗体通过Western（免疫）印迹分析检测到了该毒素。1593和2362菌株的转化体对埃及伊蚊幼虫的50%致死浓度分别为每毫升2.7×10²和5.7×10²个细胞。这种毒性水平与苏云金芽孢杆菌以色列亚种的50%致死浓度相当，但远高于球形芽孢杆菌1593和2362（每毫升4.7×10⁴个细胞）对埃及伊蚊幼虫的毒性。（摘要截短至250字）
