pEnCMV-KRAS WT-3×FLAG|BNCC356342 |BNCC

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pEnCMV-KRAS WT-3×FLAG

Culture medium	LB + 50mcg/ml ampicillin: yeast extract 5.0g, peptone 10.0g,NaCl 10.0g, distilled water 1.0L,pH 7.0. Sterilization at 121 ℃ for 15min. After sterilization, streptomycin was added when the culture medium was cooled to 40-50 ℃ with a concentration of 50mcg/ml.
Subculture procedure	1. Dissolve: After receiving the plasmid dry powder, please add 20 μl of sterile water to the bottom of the tube, dissolve the plasmid, and let it stand at room temperature for 1min; 2. Mixed (adsorption plasmid): 200 μl competent cells + plasmid DNA 5~10 μl mixed evenly and placed on ice for 30min; 3. Heat shock introduction: Let stand at 42 ℃ for 90s; 4. Shrink film hole: Ice bath for 2min; 5. Repair culture: Each tube was added with 800 μl LB liquid medium and cultured at 37 ℃ for 1h 150 r/min; 6. Screening and cultivation: Apply the appropriate volume (100 μl) of resuscitated cells on the corresponding resistant LB plate, and place it on the plate for 30min (after the agar surface must be dried), Inverted culture for 12-16h, colonies appeared. 7. Extraction: Pick the monoclonal colonies into the corresponding resistant LB liquid medium, shake culture for 12-16h, according to the test needs to extract the plasmid.
Growth conditions	LB + Ampicillin, 37 ℃ (Cloned Strain DH5α)
Storage conditions	2-8 ℃
Safety level	1
application	Scientific research
Sharing mode	Public welfare sharing