Plasmid DNA isolation

A spin-column apparatus for isolating plasmid DNA from a bacterial alkaline lysate, the apparatus comprising a mixing chamber for an alkaline lysate in fluid contact with first a filtering medium, and then a DNA binding matrix;

• wherein an immobilized concentrated P2 reagent and an immobilized P3 reagent in a dry solid or semi-solid formulation are disposed in the mixing chamber above the filtering medium, free from absorbent media;
• the P2 reagent being disposed to contact and lyse a cell suspension producing an alkaline lysate before contacting the P3 reagent, wherein the filtering media and DNA binding matrix are permeable to the alkaline lysate upon centrifugation or application of pressure;
• and wherein the DNA binding matrix is capable of substantially purifying plasmid DNA from cellular components.

A lysate filtration device for filtering a bacterial alkaline lysate, the device comprising a mixing chamber, having a top and bottom opening, a P2 reagent disposed immobilized in the mixing chamber, the P2 reagent being about a 1.5× to 15× concentrated solution containing an alcohol, a P3 reagent disposed in the mixing chamber, the P3 reagent further comprising a potassium salt, a chaotropic agent, and a solid acid, the P3 reagent being a dry formulation, a filtering media positioned in the mixing chamber, the filtering media being separate from the P2 reagent and P3 reagent and in fluid contact with the mixing chamber, the mixing chamber being configured to receive a bacterial cell suspension through the top opening, wherein contact between the P2 reagent and the bacterial cell suspension can occur in the mixing chamber, whereby contact between the P2 and the bacterial cell suspension will result in a bacterial alkaline lysate, and wherein contact between the bacterial alkaline lysate and P3 reagent can occur in the mixing chamber, whereby contact between the P3 reagent and the bacterial alkaline lysate will result in a neutralized lysate, the neutralized lysate further comprising a flocculent bacterial debris and a remaining bacterial lysate, and wherein the filtering media is for filtering the flocculent bacterial debris, from the remaining bacterial lysate.