Escherichia coli strains that allow antibiotic-free plasmid selection and maintenance by repressor titration
| Autor: | Rocky M. Cranenburgh, David J. Sherratt, Steven Geraint Williams, Julian A. J. Hanak |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
Genetic Markers
Operator (biology) Molecular Sequence Data lac operon Repressor Biology Lac repressor medicine.disease_cause Transformation Genetic Plasmid Kanamycin Escherichia coli Genetics medicine Amino Acid Sequence Cloning Molecular NAR Methods Online Plasmid preparation Base Sequence Drug Resistance Microbial Gene Expression Regulation Bacterial Chromosomes Bacterial Molecular biology Transformation (genetics) Lac Operon Plasmids |
| DOI: | 10.1093/nar/29.5.e26 |
| Popis: | We report the construction of two novel Escherichia coli strains (DH1lacdapD and DH1lacP2dapD) that facilitate the antibiotic-free selection and stable maintenance of recombinant plasmids in complex media. They contain the essential chromosomal gene, dapD, under the control of the lac operator/promoter. Unless supplemented with IPTG (which induces expression of dapD) or DAP, these cells lyse. However, when the strains are transformed with a multicopy plasmid containing the lac operator, the operator competitively titrates the LacI repressor and allows expression of dapD from the lac promoter. Thus transformants can be isolated and propagated simply by their ability to grow on any medium by repressor titration selection. No antibiotic resistance genes or other protein expressing sequences are required on the plasmid, and antibiotics are not necessary for plasmid selection, making these strains a valuable tool for therapeutic DNA and recombinant protein production. We describe the construction of these strains and demonstrate plasmid selection and maintenance by repressor titration, using the new pORT plasmid vectors designed to facilitate recombinant DNA exploitation. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|