US 10,889,793 B2
C1 substrate-fed fermentation systems and methods for producing C4 compounds
Joshua A. Silverman, Los Altos Hills, CA (US); Sol M. Resnick, Encinitas, CA (US); and Drew D. Regitsky, San Francisco, CA (US)
Assigned to CALYSTA, INC., Menlo Park, CA (US)
Filed by Calysta, Inc., Menlo Park, CA (US)
Filed on Jun. 30, 2017, as Appl. No. 15/639,120.
Application 15/639,120 is a continuation of application No. 14/434,315, granted, now 10,501,714, previously published as PCT/US2013/063650, filed on Oct. 7, 2013.
Claims priority of provisional application 61/711,104, filed on Oct. 8, 2012.
Prior Publication US 2017/0298315 A1, Oct. 19, 2017
Int. Cl. C12M 1/02 (2006.01); C12M 1/04 (2006.01); C12M 1/08 (2006.01); C12P 5/02 (2006.01); C12P 7/16 (2006.01); C12P 7/18 (2006.01); C12P 7/40 (2006.01); C12P 7/42 (2006.01); C12P 17/02 (2006.01); C12N 1/08 (2006.01); C12N 1/10 (2006.01); C12N 1/12 (2006.01); C12N 1/20 (2006.01); C12N 13/00 (2006.01); C02F 3/32 (2006.01); A01G 33/00 (2006.01); C12M 1/00 (2006.01); C12M 1/12 (2006.01)
CPC C12M 29/08 (2013.01) [C12M 23/06 (2013.01); C12M 23/34 (2013.01); C12M 27/24 (2013.01); C12M 29/06 (2013.01); C12M 41/24 (2013.01); C12P 5/02 (2013.01); C12P 7/16 (2013.01); C12P 7/18 (2013.01); C12P 7/40 (2013.01); C12P 7/42 (2013.01); C12P 17/02 (2013.01); Y02E 50/30 (2013.01); Y02P 20/582 (2015.11)] 26 Claims
OG exemplary drawing
 
1. A method for producing C4 compounds, the method comprising culturing a methanotroph comprising a heterologous polynucleotide on a C1 substrate comprising methane in a controlled culturing unit under conditions sufficient for the methanotroph comprising the heterologous polynucleotide to produce one or more of the products crotonic acid or salts thereof, n-butanol, isobutanol, 1,3-butanediol, 1,4-butanediol, and 2,3-butanediol, wherein the methanotroph is a Methylococcus capsulatus Bath, Methylomonas sp. 16a, Methylosinus trichosporium OB3b, Methylosinus sporium, Methylocystis parvus, Methylomonas methanica, Methylomonas albus, Methylobacter capsulatus, Methylobacterium organophilum, Methylomonas sp. AJ-3670, Methylomicrobium alcahphilum, Methylocella silvestris, Methylacidiphilum infernorum, Methylibium petroleiphilum, or a high growth variants thereof, wherein the culturing in the controlled culturing unit includes:
dispersing the C1 substrate comprising methane in a liquid media including at least water and one or more nutrients to form a multi-phase mixture comprising C1 substrate bubbles dispersed in the liquid media;
flowing at a first velocity and a first pressure the multi-phase mixture in one or more downward flow paths before flowing the multi-phase mixture in a number of upward flow paths each defined by a closed fluid channel, the one or more downward flow paths formed between an exterior perimeter of a number of hollow fluid conduits and an interior perimeter of a vessel at least partially surrounding the number of hollow fluid conduits, the vessel including a top and the number of hollow fluid conduits extending from the top of the vessel;
increasing the pressure of the multi-phase mixture from the first pressure to a second pressure to produce compressed C1 substrate bubbles in the multi-phase mixture while contacting the multi-phase mixture with the methanotrophs present in the one or more downward flow paths to provide a first biomass;
flowing at a second velocity the multi-phase mixture in the number of upward flow paths, each of the respective number of upward flow paths formed by an interior perimeter of each of the number of hollow fluid conduits and maintaining the pressure at or above a third pressure to maintain compressed C1 substrate bubbles in the multi-phase mixture while contacting the multi-phase mixture with the methanotrophs present in the number of upward flow paths to provide a second biomass; and
discharging the multi-phase mixture, flowing in the number of upward flow paths, from the vessel to outside the vessel through a top of the vessel without the multi-phase mixture from the upward flow path being recirculated within the vessel to the downward flow path.