Inducible T7 RNA Polymerase-mediated Multigene Expression System, pMGX
| Autor: | Kinya Hotta, Fern R. McSorley, Christopher N. Boddy, Mohamed I. Hassan |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Operon General Chemical Engineering Genetic Vectors Computational biology Biology medicine.disease_cause General Biochemistry Genetics and Molecular Biology Viral Proteins 03 medical and health sciences Synthetic biology Plasmid Escherichia coli Genetics medicine T7 RNA polymerase Gene Selectable marker Messenger RNA General Immunology and Microbiology General Neuroscience DNA-Directed RNA Polymerases 030104 developmental biology Plasmids medicine.drug |
| Zdroj: | Journal of Visualized Experiments. |
| ISSN: | 1940-087X |
| DOI: | 10.3791/55187-v |
| Popis: | Co-expression of multiple proteins is increasingly essential for synthetic biology, studying protein-protein complexes, and characterizing and harnessing biosynthetic pathways. In this manuscript, the use of a highly effective system for the construction of multigene synthetic operons under the control of an inducible T7 RNA polymerase is described. This system allows many genes to be expressed simultaneously from one plasmid. Here, a set of four related vectors, pMGX-A, pMGX-hisA, pMGX-K, and pMGX-hisK, with either the ampicillin or kanamycin resistance selectable marker (A and K) and either possessing or lacking an N-terminal hexahistidine tag (his) are disclosed. Detailed protocols for the construction of synthetic operons using this vector system are provided along with the corresponding data, showing that a pMGX-based system containing five genes can be readily constructed and used to produce all five encoded proteins in Escherichia coli. This system and protocol enables researchers to routinely express complex multi-component modules and pathways in E. coli. |
| Databáze: | OpenAIRE |
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