pCMV-T7-MCS-6×FLAG-WPRE-Neo - Basic Information

Recommended Medium	Yeast extract: 5.0g, Peptone: 10.0g, NaCl: 10.0g, pH: 7.0 (25°C)
Growth Conditions	37°C; 18-24h; Aerobic
Storage Conditions	2-8°C
Procedure	① Dissolution: Add 20 μL of sterile water to the plasmid dry powder to dissolve, let stand at room temperature for 1 min; ② Mixing: Take 200 μL of competent cells and mix with 5-10 μL of plasmid DNA, place on ice for 30 min; ③ Heat shock introduction: Let stand at 42°C for 90s; ④ Membrane pore contraction: Ice bath for 2 min; ⑤ Recovery culture: Add 800 μL of liquid Recommended Medium to each tube, culture on a shaker at 37°C for 1h (150 r/min); ⑥ Selection culture: Spread an appropriate volume (100 μL) of recovered cells onto LB plates with the corresponding antibiotic resistance, leave plates upright for 30 min (the agar surface must be dry), then invert and culture for 12-16h until colonies appear; ⑦ Extraction: Pick single monoclonal colonies into liquid Recommended Medium containing the corresponding antibiotic, culture with shaking for 12-16h, and extract the plasmid as required for the experiment.
Reference Application	pCMV-T7-MCS-6×FLAG-WPRE-Neo is a dual-promoter (CMV/T7) plasmid vector for high-level expression in mammalian cells, driving the fusion of the target gene with a 6×FLAG tag, and containing a WPRE transcriptional enhancer element and a neomycin (Neo/G418) resistance gene for stable selection.
Sharing Method	Public Welfare Sharing

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