US 11,834,651 B2
Method for producing and purifying RNA, comprising at least one step of tangential flow filtration
Andreas Funkner, Tübingen (DE); Stefanie Dorner, Tübingen (DE); Stefanie Sewing, Tübingen (DE); Johannes Kamm, Tübingen (DE); Norbert Broghammer, Tübingen (DE); Thomas Ketterer, Gomaringen (DE); and Thorsten Mutzke, Reutlingen (DE)
Assigned to CureVac Manufacturing GmbH, Tübingen (DE)
Filed by CureVac Manufacturing GmbH, Tübingen (DE)
Filed on May 16, 2023, as Appl. No. 18/198,258.
Application 18/198,258 is a continuation of application No. 17/820,242, filed on Aug. 16, 2022.
Application 17/820,242 is a continuation of application No. 17/591,978, filed on Feb. 3, 2022, granted, now 11,667,910.
Application 17/591,978 is a continuation of application No. 16/934,279, filed on Jul. 21, 2020, granted, now 11,274,293, issued on Mar. 15, 2022.
Application 16/934,279 is a continuation of application No. 15/580,092, granted, now 10,760,070, issued on Sep. 1, 2020, previously published as PCT/EP2016/062152, filed on May 30, 2016.
Claims priority of application No. PCT/EP2015/062002 (WO), filed on May 29, 2015.
Prior Publication US 2023/0287388 A1, Sep. 14, 2023
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/10 (2006.01); C12P 19/34 (2006.01); B01D 61/14 (2006.01); B01D 71/12 (2006.01); G01N 30/02 (2006.01)
CPC C12N 15/1017 (2013.01) [B01D 61/145 (2013.01); B01D 71/12 (2013.01); C12P 19/34 (2013.01); G01N 30/02 (2013.01); G01N 2030/027 (2013.01)] 45 Claims
 
1. A method for producing formulated purified RNA, comprising the steps of:
A1) providing a plasmid DNA encoding a RNA polymerase promoter sequence, a RNA coding sequence, and a restriction endonuclease cleavage site, wherein the coding sequence for said RNA is no greater than 15,000 nucleotides in length;
A2) linearizing the plasmid DNA with a restriction endonuclease to produce linearized DNA;
A3) ultrafiltering the linearized DNA by tangential flow filtration (TFF), to produce purified linearized DNA;
B1) transcribing the purified linearized DNA to produce transcribed RNA;
B2) treating the transcribed RNA with a DNase;
C1) ultrafiltering the transcribed RNA by performing at least one step of TFF using a TFF membrane cassette having a molecular weight cutoff (MWCO) of less than or equal to 500 kDa, to produce purified RNA; and
D) formulating the purified RNA to produce the formulated purified RNA.