	US 11,667,910 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 Feb. 3, 2022, as Appl. No. 17/591,978.
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 2022/0325273 A1, Oct. 13, 2022
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)]

26 Claims

1. A method for producing purified RNA, comprising the steps of:

A1) providing plasmid DNA encoding a RNA of 500 to 10000 nucleotides in length;

A2) linearizing the DNA with a restriction endonuclease to produce linearized DNA;

B) transcribing the linearized DNA to yield transcribed RNA, wherein said transcribing is in a solution comprising: nucleoside triphosphates (NTPs); T7 polymerase, spermidine, salts and a HEPES or TRIS buffer; and

C) purifying the transcribed RNA by performing at least one step of tangential flow filtration (TFF) using a TFF membrane cassette, thereby producing purified RNA, wherein the method comprises at least one step of DNA or RNA purification using chromatography.