pCMV-3×HA-mCherry-Neo - Basic Information

Recommended Medium	Yeast extract: 5.0g, Peptone: 10.0g, NaCl: 10.0g, pH: 7.0 (25℃)
Growth Conditions	37℃; 18-24h; Aerobic
Storage Conditions	2-8℃
Procedure	① Dissolution: Add 20 μL of sterile water to the plasmid dry powder to dissolve, and 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, and place on ice for 30 min; ③ Heat-shock introduction: Let stand at 42℃ for 90 s; ④ Membrane pore contraction: Ice bath for 2 min; ⑤ Recovery culture: Add 800 μL of liquid Recommended Medium to each tube, and culture on a shaker at 37℃ for 1 h (150 r/min); ⑥ Selection culture: Spread an appropriate volume (100 μL) of recovered cells onto an LB plate with the corresponding antibiotic. Leave the plate upright for 30 min (after the agar surface is completely dry), then invert and culture for 12-16 h until colonies appear; ⑦ Extraction: Pick a single monoclonal colony into liquid Recommended Medium with the corresponding antibiotic, culture with shaking for 12-16 h, and extract the plasmid as needed for the experiment.
Reference Use	pCMV-3×HA-mCherry-Neo is a plasmid vector for protein expression in mammalian cells. It uses the CMV promoter to drive the fusion expression of the 3×HA tag and mCherry red fluorescent protein, and carries a neomycin (Neo/G418) resistance gene for stable cell line selection. The vector is derived from the pmCherry-N1 backbone and uses kanamycin (Kan) as the resistance marker in bacteria.
Sharing Method	Public welfare sharing

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