Application of insecticidal protein in prevention and treatment of aphids
A technology of insecticidal protein and pests, applied in the direction of application, insecticide, biocide, etc., can solve problems such as environmental pollution, food safety, collateral damage, etc.
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[0025] Table 1 is the preparation table of SDS polyacrylamide gel.
[0026] Table 1 Preparation table of SDS polyacrylamide gel
[0027]
Embodiment
[0029] The insecticidal protein gene Cry78Aa1 (SEQ ID No.1, the amino acid sequence of the protein encoded by it is shown in SEQ ID No.2) gene in IPPBIOTSUC-B4F11 was connected to pET21b, and transformed into Escherichia coli Rosetta (DE3) , and the transformant was named Rosetta(DE3)-78A.
[0030] Extraction of insecticidal protein:
[0031] (1) Pick a single colony of Rosetta(DE3)-78A and put it in 5mL LB liquid medium (adding ampicillin and chloramphenicol), culture at 220rpm at 37°C, and wait for 2h from OD to OD 600 When the temperature is between 0.5 and 0.8, add 500 μL of 1mol / L IPTG;
[0032] (2) Adjust the temperature to 20° C., 150 rpm, and continue culturing for 12 hours to obtain a fermentation broth;
[0033] (3) Centrifuge the fermentation broth at 8000g for 10min at 4°C;
[0034] (4) Discard the supernatant, resuspend the pellet with 10mL 20mmol / L pH=8.0 Tris-HCl buffer, transfer to a 50mL centrifuge tube, and ultrasonically disrupt the cell wall (power 60%, ...
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