pLV3-CMV-4×Mito-EF1a-CopGFP-Puro - 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 lyophilized powder for dissolution, 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, then 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 at 37℃ on a shaker 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 (the agar surface must be dry), then invert and culture for 12-16 h until colonies appear; ⑦ Extraction: Pick single monoclonal colonies into liquid Recommended Medium with the corresponding antibiotic, culture with shaking for 12-16 h, and extract the plasmid as required by the experiment.
Reference Application	pLV3-CMV-4×Mito-EF1a-CopGFP-Puro is a lentiviral expression vector that uses the CMV promoter to drive the expression of the target gene labeled with 4×Mito (mitochondrial localization signal), targeting the protein of interest to mitochondria; At the same time, the EF1a promoter independently drives CopGFP green fluorescent protein as a tracking marker, and carries a Puromycin resistance gene for selection of stably transfected cell lines.
Sharing Method	Public Welfare Sharing