Efficient Disruption of Escherichia coli for Plasmid DNA Recovery in a Bead Mill
| Autor: | J. Armando Lucero-Acuña, Jaime Ortega-López, Patricia Guerrero-Germán, Armando Tejeda-Mansir, Adriana Padilla-Zamudio |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
plasmid DNA recovery Kinetics medicine.disease_cause lcsh:Technology Bead (woodworking) lcsh:Chemistry 03 medical and health sciences Plasmid dna mechanical cell lysis medicine release kinetics of pDNA General Materials Science Response surface methodology Instrumentation Escherichia coli lcsh:QH301-705.5 bead mill Fluid Flow and Transfer Processes Chromatography Chemistry lcsh:T Process Chemistry and Technology General Engineering Factorial experiment lcsh:QC1-999 Computer Science Applications 030104 developmental biology lcsh:Biology (General) lcsh:QD1-999 lcsh:TA1-2040 Yield (chemistry) Cell disruption lcsh:Engineering (General). Civil engineering (General) lcsh:Physics |
| Zdroj: | Applied Sciences; Volume 8; Issue 1; Pages: 30 Applied Sciences, Vol 8, Iss 1, p 30 (2017) |
| ISSN: | 2076-3417 |
| DOI: | 10.3390/app8010030 |
| Popis: | The release kinetics of pDNA in a bead mill was studied. Samples taken during the process were analyzed to determine total pDNA (pDNA(t)) and supercoiled pDNA (pDNA(sc)) concentration. In order to identify important variables of the process and to develop an empirical model for optimal pDNA(t) and pDNA(sc) release, a two level 23 factorial design was used with variables: mill frequency, cell concentration, and bead size. The results were analyzed by response surface methodology. The optimized conditions for pDNA(t) yield 13.26 mg/g dcw (93.41% recovery), with a mill frequency of 30 Hz, a bead size of 0.10–0.25 mm, and a cell concentration of 20 g wcw/L. However, the optimized conditions for pDNA(sc) yield 7.65 mg/g dcw (92.05% recovery), with a mill frequency of 15 Hz, a bead size of 0.10–0.25 mm, and a cell concentration of 10 g wcw/L. Cell disruption in a bead mill was proved efficient for the release of pDNA(t) and pDNA(sc) compared to the alkaline treatment. The results obtained suggest a compromise between pDNA(sc) purity and recuperation in the process development. |
| Databáze: | OpenAIRE |
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