pECMV-Nox4-m-FLAG|360113 |BNCC

Essential Information Certificate Related Products

pECMV-Nox4-m-FLAG plasmid

Subculture procedure	1. Dissolution: Upon receiving the plasmid Freeze dried pellet, add 20μl sterile water to the bottom of the tube to dissolve the plasmid. Incubate at Room temperature for 1 minute. 2. Mixing (Plasmid Adsorption): Add 200μl competent cells + 5–10μl plasmid DNA, mix thoroughly, and incubate on ice for 30 minutes. 3. Heat shock transformation: Incubate at 42°C for 90 seconds; 4. Shrink membrane wells: Ice-bathe for 2 minutes; 5. Repair culture: Add 800μl LB liquid medium to each tube; incubate at 37°C for 1 hour at 150 rpm; 6. Screening culture: Spread an appropriate volume (100 μl) of resuspended cells onto LB plates with the corresponding antibiotic. Incubate plates upright for 30 min (ensure agar surface is completely dry), then invert and incubate for 12–16 h until colonies appear. 7. Extraction: Transfer a single colony to the corresponding antibiotic-containing LB liquid medium. Incubate with shaking for 12–16 hours. Extract the plasmid as required for the experiment.
Growth conditions	37°C; 18-24h; aerobic
Storage conditions	2-8°C
Safety level	1
Sharing mode	Public welfare sharing

Product specifications: 1.0~2.0μg freeze-dried powder

Storage temperature: -20 ℃

Validity: 90 days, please convert as soon as possible

Transportation method: Transportation at room temperature, within a week

Precautions: please be sure to convert into competent cells and use them after enrichment and extraction. Before conversion, please ask the official website or the salesman to inquire about the name and concentration of the corresponding antibiotics, competent cells and culture temperature.

Use steps:

1. dissolution: after receiving the plasmid dry powder, please add 20 μL sterile water to the bottom of the tube, dissolve the plasmid, and let it stand at room temperature for 1min;

2. mixing (adsorption plasmid): 200 μl competent cells + plasmid DNA 5~10μl mix evenly and place 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: add 800 μl to each tube LB liquid medium, cultured at 37 ℃ for 1h 150 r/min;

6. screening culture: appropriate volume (100μl) resuscitated cells are coated on LB plates with corresponding resistance, placed in a plate for 30min (agar surface must be dried), cultured upside down for 12-16h, and colonies appear.

7. extraction: select monoclonal colonies into the corresponding resistant LB liquid medium, shake culture for 12-16h, and extract plasmids according to test requirements.

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