US 12,467,073 B2
Methods for producing plasmid DNA
Michael Küchler, Hamburg (DE); Christian Kaiser, Hamburg (DE); and Björn Krüger, Hamburg (DE)
Assigned to RICHTER-HELM BIOLOGICS GMBH & CO. KG, Hamburg (DE)
Appl. No. 17/916,282
Filed by RICHTER-HELM BIOLOGICS GMBH & CO. KG, Hamburg (DE)
PCT Filed Mar. 31, 2021, PCT No. PCT/EP2021/058337
§ 371(c)(1), (2) Date Sep. 30, 2022,
PCT Pub. No. WO2021/198281, PCT Pub. Date Oct. 7, 2021.
Claims priority of application No. 20167238 (EP), filed on Mar. 31, 2020.
Prior Publication US 2023/0175031 A1, Jun. 8, 2023
Int. Cl. C12N 15/10 (2006.01); C12P 19/34 (2006.01)
CPC C12P 19/34 (2013.01) [C12N 15/1006 (2013.01)] 20 Claims
OG exemplary drawing
 
1. A method of isolating covalently closed circular (ccc) DNA molecules from microbial cells containing the ccc DNA molecules, comprising the steps of:
a) contacting the microbial cells with a lysing agent and moving the composition through a tube system with a flow having a Reynolds number of at least 3000 to obtain a lysing composition;
b) incubating the lysing composition, which comprises moving the lysing composition through a tube system with a flow having a Reynolds number of at least 2100, to obtain a lysate;
c) contacting the lysate with a neutralizing solution to obtain a neutralized lysate; and
d) further processing the neutralized lysate to obtain the ccc DNA molecules.