US 11,746,321 B2
Methods of cloning nucleic acids or producing proteins in a low endotoxin organism
Matthew T Weinstock, San Diego, CA (US); Daniel G. Gibson, Carlsbad, CA (US); and Daniel Strimling, La Jolla, CA (US)
Assigned to Telesis Bio Inc., San Diego, CA (US)
Filed by Codex DNA, Inc., San Diego, CA (US)
Filed on Mar. 25, 2021, as Appl. No. 17/212,926.
Application 17/212,926 is a division of application No. 16/154,618, filed on Oct. 8, 2018, granted, now 10,968,496.
Claims priority of provisional application 62/588,755, filed on Nov. 20, 2017.
Prior Publication US 2021/0284954 A1, Sep. 16, 2021
Int. Cl. C12N 1/20 (2006.01); C12N 15/74 (2006.01); C12N 9/10 (2006.01); C12R 1/63 (2006.01)
CPC C12N 1/205 (2021.05) [C12N 9/1029 (2013.01); C12N 15/74 (2013.01); C12R 2001/63 (2021.05)] 18 Claims
 
1. A method of cloning a nucleic acid or of producing a protein comprising:
culturing a recombinant Vibrio sp. organism comprising:
an exogenous nucleic acid sequence for cloning by the organism, or an exogenous nucleic acid sequence encoding a heterologous protein or peptide for production by the organism;
a genetic modification selected from deletion, inactivation, or disruption of the lpxL gene or the lpxM gene, wherein the organism produces substantially less endotoxin compared to a corresponding organism not comprising the deletion, inactivation, or disruption and cultivated under the same conditions; and
wherein the recombinant Vibrio sp. organism exhibits a growth rate of at least 60% of the growth rate of the corresponding organism when cultivated under the same conditions;
wherein the organism has an endotoxin level of less than 50 EU/ml of purified lipopolysaccharide; and
harvesting the nucleic acid sequence cloned by the organism, or the protein or peptide produced by the organism.