Biomaterial Database

Enhanced plasmid DNA production by enzyme-controlled glucose release and an engineered Escherichia coli. 2016

Elisa A Ramírez, and Daniela Velázquez, and Alvaro R Lara

Plan de estudios en Ingeniería Biológica, Universidad Autónoma Metropolitana-Cuajimalpa, México, DF, Mexico.

OBJECTIVE To evaluate the combination of a culture medium employing glucoamylase-mediated glucose reléase from a gluco-polysaccharide and an E. coli strain engineered in its glucose transport system for improving plasmid DNA (pDNA) production. RESULTS The production of pDNA was tested using E. coli DH5α grown in shake-flasks and the recently developed VH33 Δ(recA deoR)-engineered strain, which utilizes glucose more efficiently than wild type strains. Three glucoamylase concentrations for releasing glucose from the polysaccharide carbon source were used: 1, 2 and 3 U l(-1). Both strains reached similar cell densities ranging from 5 to 8.8 g l(-1) under the different conditions. The highest pDNA yields on biomass (YpDNA/X) for both strains were obtained when 3 U enzyme l(-1)were used. Under these conditions, 35 ± 3 mgof pDNA l(-1) were produced by DH5α after 24 h of culture. Under the same conditions, the engineered strain produced 66 ± 1 mgpDNAl(-1) after 20 h. pDNA supercoiled fractionswere close to 80 % for both strains. CONCLUSIONS The pDNA concentration achieved by the engineered E. coli was 89 % higher than that of DH5α. The combination of the engineered strain and enzyme-controlled glucose release is an attractive alternative for pDNA production in shake-flasks.

UI	MeSH Term	Description	Entries
D009154	Mutation	Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations.	Mutations
D010957	Plasmids	Extrachromosomal, usually CIRCULAR DNA molecules that are self-replicating and transferable from one organism to another. They are found in a variety of bacterial, archaeal, fungal, algal, and plant species. They are used in GENETIC ENGINEERING as CLONING VECTORS.	Episomes,Episome,Plasmid
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005947	Glucose	A primary source of energy for living organisms. It is naturally occurring and is found in fruits and other parts of plants in its free state. It is used therapeutically in fluid and nutrient replacement.	Dextrose,Anhydrous Dextrose,D-Glucose,Glucose Monohydrate,Glucose, (DL)-Isomer,Glucose, (alpha-D)-Isomer,Glucose, (beta-D)-Isomer,D Glucose,Dextrose, Anhydrous,Monohydrate, Glucose
D060847	Metabolic Engineering	Methods and techniques used to genetically modify cells' biosynthetic product output and develop conditions for growing the cells as BIOREACTORS.	Engineering, Metabolic
D061249	Batch Cell Culture Techniques	Methods for cultivation of cells, usually on a large-scale, in a closed system for the purpose of producing cells or cellular products to harvest.	Batch Culture Techniques,Batch Culture,Continuous Batch Culture,Fed Batch Culture Techniques,Fed-Batch Culture,Fed-Batch Culture Techniques,Shake-Flask Culture,Batch Culture Technique,Batch Culture, Continuous,Batch Cultures,Batch Cultures, Continuous,Continuous Batch Cultures,Culture Technique, Batch,Culture Technique, Fed-Batch,Culture Techniques, Batch,Culture Techniques, Fed-Batch,Culture, Batch,Culture, Continuous Batch,Culture, Fed-Batch,Culture, Shake-Flask,Cultures, Batch,Cultures, Continuous Batch,Cultures, Fed-Batch,Cultures, Shake-Flask,Fed Batch Culture,Fed-Batch Culture Technique,Fed-Batch Cultures,Shake Flask Culture,Shake-Flask Cultures,Technique, Batch Culture,Technique, Fed-Batch Culture,Techniques, Batch Culture,Techniques, Fed-Batch Culture
D019444	Vaccines, DNA	Recombinant DNA vectors encoding antigens administered for the prevention or treatment of disease. The host cells take up the DNA, express the antigen, and present it to the immune system in a manner similar to that which would occur during natural infection. This induces humoral and cellular immune responses against the encoded antigens. The vector is called naked DNA because there is no need for complex formulations or delivery agents; the plasmid is injected in saline or other buffers.	DNA Vaccine,DNA Vaccines,Naked DNA Vaccine,Naked DNA Vaccines,Recombinant DNA Vaccine,Recombinant DNA Vaccines,Vaccines, Recombinant DNA,DNA Vaccine, Naked,DNA Vaccine, Recombinant,DNA Vaccines, Naked,DNA Vaccines, Recombinant,Vaccine, DNA,Vaccine, Naked DNA,Vaccine, Recombinant DNA,Vaccines, Naked DNA
D029968	Escherichia coli Proteins	Proteins obtained from ESCHERICHIA COLI.	E coli Proteins