pUC119|353866 |BNCC

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pUC119 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. Cooling shock: 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 revived 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 LB liquid medium with the corresponding antibiotic, shake-incubate for 12-16 hours, and extract the plasmid as required for the experiment.
Growth conditions	LB + Ampicillin, 37°C (Cloning strain DH5α)
Storage conditions	2-8°C
Safety level	1
Sharing mode	Public welfare sharing

Product format: freeze dried, 1.0-2.0ug
Validity period: 90 days
Storage temperature: -20℃
Shipping: at ambient temperature,within one week

Notes: The plasmids shall be transformed into competent cells, and then use after amplification and extraction. Before transformation, please contact technicians or visit the website for the information about the name and concentration of the antibiotics, competent cells and culture temperature.

Handling procedures:

1 Dissolving: upon receipt of the freeze dried plasmid, add 20ul of sterile water to the bottom of the ampoule, dissolve the pellets, and sit at room temperature for 1min;

2 Mixture (adsorption of plasmids ): mix 200ul competent cells and 5-10ul of plasmid DNA evenly and place it on ice for 30min;

3 Heat shock: sit at 42 ℃ for 90s

4 Shrink the membrane pore: ice bath for 2min

5 Repair cultivation: add 800ul of LB liquid medium to each tube, culture at 37 ℃for 1 hour,
150 r / min;

6 Screening cultivation: distribute appropriate (100ul) recovered cells to the LB plate with corresponding antibiotics, place the plate uprightly for 30min (the agar surface must be dried), sit the plate upside down for 12-16h, and colonies appear.

7 Extraction: pick the monoclonal colonies into the LB liquid medium with corresponding antibiotics, shake for 12-16h, and extract the plasmid according to the needs.

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