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Modified Escherichia coli B (BL21), a superior producer of plasmid DNA compared with Escherichia coli K (DH5alpha).
Biotechnol Bioeng. 2008 Nov 01; 101(4):831-6.BB

Abstract

Plasmid DNA (pDNA) is an emerging experimental vaccine, produced in E. coli, initially targeted for viral diseases. Unlike traditional protein vaccines whose average dose is micrograms, the average dose of pDNA is on the scale of milligrams. Production yields are, therefore, important for the future development of this vaccine. The E. coli strains currently used for pDNA production, JM109 and DH5alpha, are both suitable for production of stable pDNA due to the deletion of recA and endA, however, these two E. coli K strains are sensitive to growth conditions such as high glucose concentration. On the other hand E. coli BL21 is less sensitive to growth conditions than E. coli JM109 or DH5alpha, this strain grows to higher densities and due to its active glyoxylate shunt and anaplerotic pathways is not sensitive to high glucose concentration. This strain is used for recombinant protein production but not for pDNA production because of its inability to produce stable pDNA. To adapt E. coli BL21 for stable pDNA production, the strain was mutated by deleting both recA and endA, and a proper growth and production strategy was developed. Production values, reaching 2 g/L were obtained using glucose as a carbon source. The produced plasmid, which was constructed for HIV clinical study, was found to have identical properties to the plasmid currently produced by E. coli DH5alpha.

Authors+Show Affiliations

Biotechnology Core Laboratory, NIDDK NIH Bethesda, Bldg 14A Room 173, Maryland 20892, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Research Support, N.I.H., Intramural

Language

eng

PubMed ID

18814292

Citation

Phue, Je-Nie, et al. "Modified Escherichia Coli B (BL21), a Superior Producer of Plasmid DNA Compared With Escherichia Coli K (DH5alpha)." Biotechnology and Bioengineering, vol. 101, no. 4, 2008, pp. 831-6.
Phue JN, Lee SJ, Trinh L, et al. Modified Escherichia coli B (BL21), a superior producer of plasmid DNA compared with Escherichia coli K (DH5alpha). Biotechnol Bioeng. 2008;101(4):831-6.
Phue, J. N., Lee, S. J., Trinh, L., & Shiloach, J. (2008). Modified Escherichia coli B (BL21), a superior producer of plasmid DNA compared with Escherichia coli K (DH5alpha). Biotechnology and Bioengineering, 101(4), 831-6. https://doi.org/10.1002/bit.21973
Phue JN, et al. Modified Escherichia Coli B (BL21), a Superior Producer of Plasmid DNA Compared With Escherichia Coli K (DH5alpha). Biotechnol Bioeng. 2008 Nov 1;101(4):831-6. PubMed PMID: 18814292.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Modified Escherichia coli B (BL21), a superior producer of plasmid DNA compared with Escherichia coli K (DH5alpha). AU - Phue,Je-Nie, AU - Lee,Sang Jun, AU - Trinh,Loc, AU - Shiloach,Joseph, PY - 2008/9/25/pubmed PY - 2008/11/19/medline PY - 2008/9/25/entrez SP - 831 EP - 6 JF - Biotechnology and bioengineering JO - Biotechnol. Bioeng. VL - 101 IS - 4 N2 - Plasmid DNA (pDNA) is an emerging experimental vaccine, produced in E. coli, initially targeted for viral diseases. Unlike traditional protein vaccines whose average dose is micrograms, the average dose of pDNA is on the scale of milligrams. Production yields are, therefore, important for the future development of this vaccine. The E. coli strains currently used for pDNA production, JM109 and DH5alpha, are both suitable for production of stable pDNA due to the deletion of recA and endA, however, these two E. coli K strains are sensitive to growth conditions such as high glucose concentration. On the other hand E. coli BL21 is less sensitive to growth conditions than E. coli JM109 or DH5alpha, this strain grows to higher densities and due to its active glyoxylate shunt and anaplerotic pathways is not sensitive to high glucose concentration. This strain is used for recombinant protein production but not for pDNA production because of its inability to produce stable pDNA. To adapt E. coli BL21 for stable pDNA production, the strain was mutated by deleting both recA and endA, and a proper growth and production strategy was developed. Production values, reaching 2 g/L were obtained using glucose as a carbon source. The produced plasmid, which was constructed for HIV clinical study, was found to have identical properties to the plasmid currently produced by E. coli DH5alpha. SN - 1097-0290 UR - https://www.unboundmedicine.com/medline/citation/18814292/Modified_Escherichia_coli_B__BL21__a_superior_producer_of_plasmid_DNA_compared_with_Escherichia_coli_K__DH5alpha__ L2 - https://doi.org/10.1002/bit.21973 DB - PRIME DP - Unbound Medicine ER -