Inducible gene expression composition for using eukaryotic pol-2 promoter-driven transcription in prokaryotes and the applications thereof
Eukaryotic protein-coding messenger RNAs and non-coding microRNAs are naturally transcribed by type II RNA polymerases (pol-2) but not prokaryotic RNA polymerases. As a result, current eukaryotic RNA and protein production is performed either using eukaryotic pol-2 promoters in hybridomas or mammalian cells or using prokaryotic promoters in bacterial cells. However, because prokaryotic RNA transcription tends to be error-prone, frequent mutation is a big problem. Also, growing hybridomas or mammalian cells is relatively laborious and costly. To overcome these problems, the present invention provides a novel inducible composition and method for producing eukaryotic RNAs and/or their related peptides/proteins directly using eukaryotic pol-2 promoter-driven gene expression in fast growing bacteria, without the need of changing to prokaryotic promoters or growing hybridomas/mammalian cells. The RNAs and peptides/proteins so obtained can be used to develop drugs, cure diseases, treat tumors/cancers, produce pluripotent stem (iPS) cells, enhance wound healing, and make foods.
1. A method for inducing or enhancing eukaryotic EF1alpha promoter-driven gene expression in prokaryotes, comprising:
contacting a chemical agent, containing a structure of 3-morpholinopropane-1-sulfonic acid (MOPS) with said prokaryotes containing said eukaryotic EF1alpha promoter-driven gene, wherein said eukaryotic EF1alpha promoter-driven gene encodes at least a hairpin-like RNA structure in its 5′-untranslated region (5′-UTR).
2. The method as defined in claim 1 , further comprising mixing said chemical agent with glycerin to enhance the sufficiency of eukaryotic EF1alpha promoter-driven gene expression.
3. The method as defined in claim 1 , wherein said method is performed in a bacterial culturing medium.
4. The method as defined in claim 3 , wherein said chemical agent has a v/v concentration of 0.01% to 1% in said bacterial culturing medium.
5. The method as defined in claim 3 , wherein said bacterial culturing medium is Luria-Bertani (LB) broth.
6. The method as defined in claim 1 , wherein said eukaryotic EF1alpha promoter-driven gene encodes at least a protein/peptide and non-coding RNA, which is useful for pharmaceutical or therapeutic application.
7. The method as defined in claim 6 , wherein said non-coding RNA is small hairpin RNA useful for pharmaceutical or therapeutic application.
8. The method as defined in claim 6 , wherein said non-coding RNA is a miR-302 homologue.
9. The method as defined in claim 6 , wherein said protein or peptide is an enzyme useful for pharmaceutical or therapeutic application.
10. The method as defined in claim 1 , wherein said prokaryotes are bacterial cells.
11. The method as defined in claim 10 , wherein said bacterial prokaryotic cells are Escherichia coli ( E. coli ).
12. The method as defined in claim 1 , wherein said eukaryotic EF1alpha promoter-driven gene is located in a plasmid vector.
13. The method as defined in claim 12 , wherein said plasmid vector is pLenti-EF1alpha-RGFP-miR302.