Applications such as transfection normally require a large quantity of plasmid that meets a number of stringent criteria, including high purity and a high percentage of super coiling. The most common difficulty for large scale plasmid isolation is the contamination of endotoxins which can have a negative effect on downstream applications such as transfection.

Principle

The Endo Free Plasmid Miniprep Kit combines the power of HiBind^® technology with the time-tested consistency of alkaline-SDS lysis of bacterial cells to deliver high quality DNA with lower levels of endotoxins. Omega Bio-Tek's HiBind^® spin columns facilitate the binding, washing, and elution steps thus enabling multiple samples to be simultaneously processed.

Procedure

Following lysis, the cleared lysate is extracted with ETR Buffer which will almost completely remove the endotoxin. Then the lysate is applied to the HiBind^® Mini columns, plasmid DNA is bound to the silica membrane and contaminants are removed with a simple wash step. Yields vary according to plasmid copy number, E.coli strain, and conditions of growth, but up to 30 - 50 mg of high quality plasmid can be purified from 5 ml culture overnight.

Applications

DNA purified using this kit can be used for the following applications :

• Automated fluorescent DNA sequencing
• Transfection of Mammalian cells
• Restriction Endonuclease Digestion