pRSETA|358632 |BNCC

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pRSETA

Culture medium	BNCC361029
Description	LB Medium with 50μg/mL Ampicillin
Composition	Yeast Extract:5.0g, Peptone:10.0g, NaCl:10.0g, pH:7.0(25℃)
Growth conditions	LB + Ampicillin, 37°C (Cloning strain DH5α)
Subculture procedure	1. Dissolution: Upon receiving the plasmid powder, add 20 μl sterile water to the bottom of the tube to dissolve the plasmid. Incubate at Room temperature for 1 minute; 2. Mix (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 Pores: Ice bath 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 the 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 the corresponding antibiotic-supplemented LB liquid medium. Incubate with shaking for 12–16 hours. Extract the plasmid as required for the experiment.
Storage conditions	2-8°C
Sharing mode	Public welfare sharing