High-Yield Plasmid DNA Fermentation Process
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• 2 g/liter and more, plasmid DNA production
• Fermentation process for fed-batch modes of production
• Animal Product Free (APF) upon request
• Scalable custom fermentation
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Plasmid DNA has traditionally been grown, either inE. coli shake flask cultures (typically, 1-10 mg/L), or in batch cultures, or fed-batch fermentation cultures (typically, 50-250mg/L). However, the advent of DNA vaccines and gene medicines has created a need for a robust fermentation process with dramatically improved yield and most economical cost of goods.
Nature Technology Corp. (NTC) has created the HyperGRO™ fed-batch fermentation process to address this need, resulting in record yields exceeding 2.2g/L (Fig.1). The process utilizes a semi-defined HyperGRO™ media, which can be obtained directly from NTC. It’s use in your facility requires the ability to perform exponentially fed fermentations (NTC will provide technical assistance to your engineering staff) (Fig.2).
NTC also offers a service to produce your plasmids in our facility using the HyperGRO™ process, or we can provide you with the E. coli cell paste for purification of plasmids in your facility, after growth at NTC. This allows you to use your current downstream process, without the need for special engineering skills in the upstream process (Fig. 3).
Finally, NTC offers site-licenses for contract manufacturing facilities, or for in-house pharmaceutical production. (See HyperGro™ licensing)
Figure 1. Typical yields (mg/L, left scale) for shake flask cultures vs. HyperGRO™ batch and fed batch or fed batch modes of fermentation.
The unique components of the HyperGRO™ process contribute significantly to the plasmid production requirements of E. coli. HyperGRO™ consistently outperforms published methods, and is highly compatible with plasmid purification processes.
The HyperGRO™ process a ready-to-use product, composed of a proprietary blend of amines, trace minerals, vitamins and carbon sources optimized for maximal plasmid specific yield.
Yield may vary based on the plasmid/host combination used. Observed yields vary from approximately 100-250mg/l (SB) or 150-2,000mg/l (SFB) with average plasmids having pUC or equivalent origins of replication, grown in E. coli host DH5a. For maximizing yields, we recomend the combination of an optimizing pUC plasmid backbone (such as NTC7264) and DH5a.
Figure 2. Fed-batch HyperGRO™ (Patents Pending) fermentation using NTC7264 backbone with 30-42C temperature shift, yield 1,500mg/L (left scle, mg/L).
Figure 3. HyperGRO™ Process. Do it yourself with the HyperGRO™ process or let us provide you with the highly concentrated, E. coli Cell Paste containing your plasmid DNA.
Further information and data regarding the HyperGRO™ process:
1) Fermentation poster, ASGT 2006:
http://natx.com/PDFs/FermentationASGT2006.pdf
2) Carnes AE, Hodgson CP, Williams, JA. (2006)
Inducible Escherichia coli fermentation for increased plasmid DNA production.
Biotechnol Appl Biochem. 45:155-166. "Mounted on the internet with permissiion from Portland Press."
3) Carnes, AE, Williams, JA (2004), PROCESS FOR PLASMID DNA FERMENTATION, PCT/US2005/29238; WO2006/023546;
4) Williams, JA (2005)
COMPOSITIONS AND PROCESSES FOR IMPROVED PLASMID DNA PRODUCTION;
PCT/US2006/03148, WO 2006/081529
NTC Contact:
Clague Hodgson, Ph.D.,
Tel: 402 472-6530
Email: hodgson@natx.com
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