苏云金芽胞杆菌杀蚊基因在鱼腥藻中克隆和表达的初步研究
PRELIMINARY STUDIES ON CLONING AND EXPRESSION OF BACILLUS THURINGIENSIS CRY11A GENE IN ANABAENA
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摘要: 将苏云金芽胞杆菌以色列亚种的杀蚊晶体蛋白基因cry11A亚克隆到大肠杆菌-蓝藻的穿梭质粒载体pRL25C,然后用三亲本杂交的方法将重组质粒转移到一种具有固氮能力且可被蚊幼虫吞食的鱼腥藻(Anabaena)PCC7120中。Southernblot及Westernblot分析表明cry11A基因在鱼腥藻PCC7120中得以克隆和表达,但生物测定未能检测到转基因鱼腥藻对库蚊(Culex)的毒性,可能是因为带有苏云金芽胞杆菌自身启动子的cry11A基因在鱼腥藻PCC7120中表达量不够高的缘故。Abstract: The mosquito specific cry 11A gene of Bacillus thuringiensis subsp.israelensis was cut out from pGEM1 with BamHI and ligated into the BamHI site of a E. coli-cyanobacteriun shuttle vector pRL25C. The recombinant plasmid was transformed into E. coli HB101 which bears plasmid pRL528, a helper plasmid carrying mob and genes for Eco47II and AvaI methylase. Then three parents: E. coli HB101 (bearing the recombinant plasmid and helper plasmid pRL528), E. coli HB101 (bearing RP4) and Anabaena PCC7120 (a filamentous cyanobacterium which can fix nitrogen and can be fed by mosquito larvas in water) were put together to do triparent matings. Both Southern blot and Western blot demonstrated that the recombinant plasmid existed and expressed in the positive conjugant of Anabaena PCC7120. But the expression of the cry 11A gene with its original promoter in transgenic Anabaena PCC7120 was too low to kill mosqinto larvae.
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Keywords:
- Bacillus thuringiensis /
- Mosquito-killing gene /
- Anabaena PCC7120
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