Sigma Life Science is committed to bringing you Greener Alternative Products, which adhere to one or more of The 12 Principles of Greener Chemistry. This product has Inherently Safer Chemistry, compared to the standard use of phenol and chloroform to perform DNA extractions.
The GenElute HP Select Plasmid Gigaprep Kit offers a simple, rapid, and cost effective method for isolating endotoxin-free plasmid DNA from recombinant E. coli cultures. Up to 15 mg of high copy plasmid DNA with ≤0.1 endotoxin unit/μg plasmid DNA can be recovered from E. coli cultures in 1.2-2.5 L Luria Broth (LB) medium or 600 mL-1.2 L of Terrific Broth (TB) medium in less than 2 hours after harvesting the cells. Note that the actual yield depends on the strain, the plasmid, and the culture medium used.
An overnight recombinant E. coli culture is harvested by centrifugation and subjected to a modified alkaline-SDS lysis. The lysate is cleared by filtration followed by the addition of a binding solution optimized for endotoxin-free plasmid preparations. The plasmid DNA is then captured onto a silica membrane in the presence of high salts while endotoxins are prevented from adsorbing to the membrane. Contaminants are removed with three wash steps. Finally, the bound DNA is eluted in endotoxin-free water.
The DNA is ready for immediate use in applications such as restriction digest, cloning, sequencing, PCR, and transfection of endotoxin-sensitive cell lines.