US 12,344,873 B2
Anaerobic preconditioning of cells for improved in vitro protein production
Denis Tamiev, Ames, IA (US); and Nigel Forest Reuel, Ames, IA (US)
Assigned to Iowa State University Research Foundation, Inc., Ames, IA (US)
Filed by Iowa State University Research Foundation, Inc., Ames, IA (US)
Filed on Mar. 8, 2022, as Appl. No. 17/689,586.
Claims priority of provisional application 63/158,044, filed on Mar. 8, 2021.
Prior Publication US 2022/0282286 A1, Sep. 8, 2022
Int. Cl. C12P 1/04 (2006.01); C12N 1/20 (2006.01); C12P 21/00 (2006.01); C12R 1/19 (2006.01)
CPC C12P 1/04 (2013.01) [C12N 1/205 (2021.05); C12P 21/00 (2013.01); C12N 2500/02 (2013.01); C12N 2500/05 (2013.01); C12N 2500/34 (2013.01); C12N 2500/46 (2013.01); C12R 2001/19 (2021.05)] 14 Claims
 
1. A cell-free protein expression method comprising:
exposing a microorganism to substantially anaerobic growth conditions to produce a conditioned microorganism;
lysing the conditioned microorganism to produce a lysate; and
combining the lysate with a nucleic acid and producing a protein of interest, a combination of proteins of interest, a molecule, or a mixture thereof from the lysate,
wherein exposing the microorganism to substantially anaerobic conditions comprises exposing the microorganism to an atmosphere comprising about 90 to 99% N2 and about 1 to 10% CO2 for a time in a range of from about 5 hours to about 48 hours and the microorganism is bacteria selected from comprises-Streptococcus bacteria, Lactobacillus bacteria, Bacillus bacteria, Escherichia bacteria, Salmonella bacteria, Neisseria bacteria, Acetobactor bacteria, Arthrobacter bacteria, Aspergillus bacteria, Bifdobacterium bacteria, Corynebacterium bacteria, Pseudomanas bacteria, or a mixture thereof.