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Development of a continuous microbial fermentation system using reduced genome E. coli that undergo programmed autolysis and generate nuclease post-production for the manufacture of biopharmaceuticals

Award Number: R44GM161364
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 09/15/2025
PERIOD OF PERFORMANCE END DATE: 08/31/2027

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Development of a continuous microbial fermentation system using reduced genome E. coli that undergo programmed autolysis and generate nuclease post-production for the manufacture of biopharmaceuticals - Continuous bioprocessing that brings with it the advantages of lower ultimate cost, a smaller equipment footprint and energy efficiency, is beginning to be embraced by the biopharmaceutical industry. Long term continuous fermentation requires a high level of genetic and metabolic stability. Scarab Genomics proprietary reduced genome E. coli strains provide the requisite genetic and metabolic stability for such long term continuous fermentations. Scarab Genomics proposes to develop the first fully continuous platform for the production and purification of recombinant proteins and plasmid DNA (pDNA) by replacing homogenization of bacteria with a custom autolysis system controlled by a temperature sensitive promoter. Distinctive features of Scarab’s Clean Genome bacteria such as the absence of all cryptic prophage make it uniquely suited for tightly regulated lysis. Having attained proof-of-concept in small scale culture, this application proposes to optimize this lysis system in high density (>200 OD600) continuous fermentation (C-Flow). To further facilitate the development of a continuous system, the lysis operon will also drive the exonuclease bensonase so that as lysis occurs, the large amount of nucleic acid that hinders downstream processing will be removed. To enable a lysis system for plasmid DNA (pDNA) purification, bensonase in the lysis operon will be replaced by the lambda exonuclease which digests genomic DNA but not pDNA thereby providing an alternate system for the production of pDNA. Once optimized, this lysis and nucleic acid digestion system will be used to express two commercially relevant test proteins. The first will be insulin because Scarab sees the possibility of C-Flow as a vehicle for the production of less expensive insulin and insulin related compounds to meet the objectives of the US government and the WHO. The second will be a single chain antibody (scFv) given the numerous applications in diagnostics and therapeutics for single chain antibodies and antibody fragments. To prepare C-Flow for market, the application also proposes to develop upgrades to the current system including oxygen infusion of medium using microbubble delivery and a microbubble separation chamber for the removal of cell debris. These innovations will result in the first continuous fermentation platform in E. coli for the production of biopharmaceuticals.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2025 ( Subtotal = $937,537 )
2025	2025	SCARAB GENOMICS LLC	1202 ANN ST	MADISON	WI	53713	DANE	USA	Biomedical Research and Research Training	000	1	9/11/2025	NEW	$937,537
														Subtotal = $937,537

Grand Total All Awards = $937,537

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Development of a continuous microbial fermentation system using reduced genome E. coli that undergo programmed autolysis and generate nuclease post-production for the manufacture of biopharmaceuticals

Award Number: R44GM161364

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

PERIOD OF PERFORMANCE START DATE: 09/15/2025

PERIOD OF PERFORMANCE END DATE: 08/31/2027

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