pCold IV|394198 |BNCC

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pCold IV

Growth conditions	37°C; 18–24 h; aerobic
Subculture procedure	① Dissolution: Add 20 μL of sterile water to the plasmid powder to dissolve it, then let stand at Room temperature for 1 min; ② Mixing: Mix 200 μL of competent cells with 5–10 μL of plasmid DNA, then incubate on ice for 30 min; ③ Heat shock transformation: Incubate at 42°C for 90 s; ④ Punch holes in the shrink wrap: Incubate on ice for 2 min; ⑤ Recovery culture: Add 800 μL of liquid medium to each tube and incubate on a shaking incubator at 37°C for 1 h (150 r/min); ⑥ Screening culture: Spread an appropriate volume (100 μl) of resuspended cells onto the corresponding resistance LB agar plate; incubate the Agar plate upright for 30 min (ensuring the agar surface is completely dry), then incubate inverted for 12–16 h until colonies appear; ⑦ Isolation: Transfer single colonies to the corresponding antibiotic-containing LB liquid medium, incubate with shaking for 12–16 h, and extract the plasmid as required for the experiment.
Storage conditions	2–8°C
Sharing mode	Public welfare sharing