METHOD OF SEPARATING EXTRA-CHROMOSOMAL DNA FROM OTHER CELLULAR COMPONENTS

摘要

THE INVENTION RELATES TO A METHOD OF SEPARATING EXTRA-CHROMOSOMAL DNA FROM RNA. IT ALSO RELATES TO DNA PRODUCED BY THE METHOD AND PHARMACEUTICALS DERIVED FROM SUCH DNA, FOR EXAMPLE, DNA VACCINES. THE METHOD COMPRISES SEPARATING EXTRA-CHROMOSOMAL DNA FROM RNA WITHOUT FIRST DIGESTING THE RNA, AND COMPRISES:i) LYSING CELLS COMPRISING EXTRACHROMOSOMAL DNA TO FROM A LYSATE;@ii) CLARIFYING THE LYSATE; AND @ii) SUBJECTING THE CLARIFIED LYSATE TO TANGENTIAL FLOW ULTRA FILTRATION UNDER CONDITIONS, WHICH ALLOW A SUBSTANTIAL AMOUNT OF THE RNA TO PASS THROUGH A MEMBRANE WHILST THE EXTRA-CHROMOSOMAL DNA IS RETAINED. THE PREFERRED CONDITIONS INCLUDE THE USE OF A DIAFILTRATION BUFFER OF LOW CONDUCTIVITY AND DIAFILTRATION VOLUMES OF OVER 10 VOLUME EQUIVALENTS.