The E-Z 96^® BAC/PAC DNA Isolation kit provides a fast, reliable, high throughput method for the purification of BAC or PAC DNA. It is based on a modified alkaline lysis of bacterial cells, followed by the clearing of the lysate with an E-Z 96 ^® FastFilter^® Plate. The BAC or PAC DNA is further purified and concentrated by isopropanol precipitation.

Procedure

PAC, BAC, or P1 cultures are grown in YT medium in a 96-well deep well plate. Cells are collected by centrifugation, and re-suspended in Buffer T1/RNase A Buffer T2 is added to the re-suspended cells resulting in a cleared lysate. T3 is added and reaction is mixed gently by inverting and rotating the plate until a flocculent white precipitate forms. Carefully transfer the cell lysate to a E-Z 96^® Filter plate kept on top of a 2 ml collection plate and centrifuge to filter it. Add glycogen and room temperature isopropanol to the cleared cell lysate, mix by vortexing and centrifuge to pellet the DNA. Wash the DNA pellet with 70 % ethanol and air dry the pellet. Re-dissolve the DNA pellet in TE or molecular grade water by incubating overnight at room temperature.