The NucleoSpin® Plasmid Transfection-grade procedure is a modified version of the Birnboim and Doly alkaline lysis plasmid miniprep protocol. Pelleted bacteria are resuspended in Buffer A1 and plasmid DNA is liberated from the cells by SDS/alkaline Lysis Buffer A2. Buffer A3 neutralizes the lysate, precipitates genomic DNA, proteins and cell debris, and creates appropriate conditions for binding of plasmid DNA to the silica membrane.

The crude lysate is cleared by filtration and brought into contact with a silica membrane where plasmid DNA binds to the surface. Endotoxins and proteins are removed by the innovative Detoxification Buffer ERB. Further contaminations such as salts are removed with ethanolic Buffer AQ while traces of ethanol are removed by  vacuum. Pure plasmid DNA is eluted under low ionic strength conditions with slightly alkaline Buffer AE (5 mM Tris/HCl, pH 8.5) and is ready for any common downstream application including transfection (research use only).

Required hardware
 

Vacuum processing

Features

Applications

• Manual and automated isolation of high-copy plasmids
• Typical downstream applications: transfection of common cell lines, cell-based assays, screenings

* The NucleoSpin® 96 Plasmid Transfection-grade Core Kit provides the buffers, RNase A, NucleoSpin® Plasmid Filter Plate, and NucleoSpin® Plasmid Binding Plate only. Accessory components (e.g., culture plate, elution plate, and Wash Buffer AW) are not provided in the core kits but can individually be selected from a variety of suitable accessories (see ordering information). This allows highest flexibility for the user.