Bt (Bacillus thuringiensis) protein with insecticidal activity to lymantria dispar and application of Bt protein
A kind of insecticidal activity, gypsy moth technology, applied in the field of Bt protein
Active Publication Date: 2015-07-01
INST OF PLANT PROTECTION CHINESE ACAD OF AGRI SCI
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Problems solved by technology
The synergistic effect between Cry proteins has been confirmed in a variety of agricultural and forestry pests, but there is no synergistic research on the Bt protein of the gypsy moth. Therefore, on the basis of screening out the highly virulent Bt protein, explore several The synergistic virulence effect of the seed protein on the gypsy moth will further improve the control effect on the gypsy moth, and will provide an excellent insecticidal gene combination for the construction of engineering bacteria and the cultivation of transgenic insect-resistant crops, delaying the emergence of resistance
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Embodiment 1
[0041] 1. Protein preparation and quantification
[0042] 1) Expression of Cry1Bb, Cry1Be, Cry9Ee, Vip3Ag, Vip3Aj, Vip3Aa proteins in Escherichia coli
[0043] 37°C, 230rpm, activate the strains for 12h; inoculate 1% in 200mL LB medium (add 1 / 1000 corresponding antibiotics), cultivate at 37°C, 220rpm until the OD600 value is 0.5; add the inducer IPTG, the final concentration is 0.5mM, 150rpm, 30°C induction for 12 hours; 4°C, centrifuge at 12000rpm for 10min to collect the cells, suspend in 20mmol / L Tris-HCl (pH 8.0); ultrasonically disrupt the cells, power 80%, sonicate for 10min (over 3s and stop for 5s); 4°C , centrifuge at 12000rpm for 10min; collect the supernatant and precipitate for detection respectively, and suspend the precipitate with 20mmol / L Tris-HCl (pH 8.0); analyze by SDS-PAGE electrophoresis, see figure 1 .
[0044] 2) Expression of Cry2Ab, Cry1Ab, Cry1Ca, Cry1Ac, Cry1Ah proteins in Bacillus thuringiensis
[0045] 30°C, 230rpm, activate the strain for 12h; ...
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Abstract
The invention provides a Bt (Bacillus thuringiensis) protein with insecticidal activity to lymantria dispar and an application of the Bt protein. The protein comes from Bt and is selected from at least one of Cry1Ac protein, Cry2Ab protein, Cry1Ca protein or Vip3Aj protein. Besides, the invention also provides a Bt protein combination obtained by virtue of paired proportioning of the Bt protein. The protein with insecticidal activity to lymantria dispar provides a new material for effective control of lymantria dispar, and meanwhile, the exploration of a new gene provides an effective gene source for culturing a transgenic plant and producing a genetic engineering insecticide. Moreover, the Bt protein combination with high-toxicity to lymantria dispar not only can improve the insecticidal activity but also can effectively solve and inhibit the problems that the insecticidal spectrum is narrow, harmful insects generate resistance easily and the like in application of Bt univalent genes.
Description
technical field [0001] The invention relates to the field of biotechnology, in particular to several Bt proteins with insecticidal activity against gypsy moths, Bt protein combinations and applications thereof. Background technique [0002] The gypsy moth Lymantria dispar (Linnaeus) has the characteristics of wide distribution, miscellaneous feeding habits, serious damage, and downwind migration of larvae. It is a worldwide agricultural and forestry pest. It originated in Eurasia, and is currently mainly distributed in Asia, Europe and North America. It has caused large-scale outbreaks in the temperate zone of the whole north. Gypsy moths are distributed in almost all provinces in China. The larvae feed on leaves and harm a variety of coniferous and broad-leaved tree species and fruit trees. There are more than 500 hosts, especially poplar, willow, birch, maple, oak, etc. Even harm apples, pears, citrus and other crops. It is a long-term and arduous task to effectively con...
Claims
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Login to View More IPC IPC(8): C07K14/325A01N47/44A01P7/04
CPCA01N47/44C07K14/325C12N15/8286
Inventor 张杰杨帆陈敏束长龙耿丽丽宋福平彭琦梁影屏
Owner INST OF PLANT PROTECTION CHINESE ACAD OF AGRI SCI