Abstract: The invention relates to a process for isolating and purifying a polynucleotide on a manufacturing scale which uses a chromatographic separation process comprising a combination of two different chromatographic steps selected from hydrophobic interaction chromatography, polar interaction chromatography and anion exchange chromatography. In at least one of the two steps the chromatographic support is a porous monolithic bed. The invention also relates to a chromatographic device and its use for isolating and purifying a polynucleotide of interest, in particular plasmid DNA, on manufacturing scale.