p4412-EGFP - Basic Information

Recommended Medium	Yeast extract: 5.0 g, Peptone: 10.0 g, NaCl: 10.0 g, pH: 7.0 (25°C)
Growth Conditions	Basal Recommended Medium (LB): Yeast extract 5.0 g, Peptone 10.0 g, NaCl 10.0 g, Distilled water 1.0 L, pH 7.0. Sterilize at 121°C for 15 min. (After the Recommended Medium has been sterilized and cooled to 48°C, add ampicillin to a final concentration of 100 μg/mL.)
Storage Conditions	2-8°C
Procedure	① Dissolution: Add 20 μL of sterile water to the plasmid dry powder to dissolve it, and let it 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°C 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°C on a shaker for 1 h (150 r/min); ⑥ Selection culture: Spread an appropriate volume (100 μL) of recovered cells onto LB plates with the corresponding antibiotic. Leave the plates 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 LB liquid Recommended Medium with the corresponding antibiotic, culture with shaking for 12-16 h, and extract the plasmid according to experimental needs.
Intended Use	p4412-EGFP is an Escherichia coli-Bacillus shuttle expression vector that uses the strong constitutive promoter of Bacillus p4412 to drive expression of EGFP (enhanced green fluorescent protein), for constitutive green fluorescent labeling without induction in various Bacillus species such as Bacillus subtilis and Bacillus amyloliquefaciens.
Sharing Method	Public welfare sharing