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 composition for regulating eukaryotic elongation factor-1 alpha (EF1alpha) promoter-driven gene expression in prokaryotes, comprising:
(a) at least a chemical agent capable of inducing or enhancing eukaryotic EF1alpha promoter-driven gene expression in prokaryotic cells, wherein said chemical agent containing a structure of 3-morpholinopropane-1-sulfonic acid (MOPS); and
(b) a plurality of prokaryotic cells, said plurality of prokaryotic cells containing at least a gene mediated by a eukaryotic EF1alpha promoter-driven expression mechanism and wherein said gene encodes at least a hairpin-like structure in its 5′-untranslated region (5′-UTR) for being expressed in prokaryotes;
wherein (a) and (b) are mixed together under a condition to induce the eukaryotic EF1alpha promoter-driven gene expression of said gene in said prokaryotic cells; wherein said condition is in Luria-Bertani (LB) broth.
2. The composition as defined in claim 1 , wherein each one of said prokaryotic cells has at least an inducible gene expression composition.
3. The composition as defined in claim 2 , wherein said gene is located in a plasmid vector containing at least a eukaryotic EF1alpha promoter.
4. The composition as defined in claim 1 , wherein said gene encodes at least a non-coding RNA and a protein-coding RNA.
5. The composition as defined in claim 4 , wherein said non-coding RNA is a small hairpin RNA.
6. The composition as defined in claim 4 , wherein said protein-coding RNA further encodes a red fluorescent protein (RGFP).
7. The composition as defined in claim 4 , wherein said protein-coding RNA contains 30% to 100% homology to a cDNA sequence of a eukaryotic gene.
8. The composition as defined in claim 4 , wherein said non-coding RNA or said protein-coding RNA is isolatable from said prokaryotes.
9. The composition as defined in claim 1 , wherein said gene encodes at least a protein.
10. The composition as defined in claim 4 , wherein a protein transcribed from said protein-coding RNA contains at least a peptide sequence.
11. The composition as defined in claim 10 , wherein said protein or said peptide is isolatable from said prokaryotes.
12. The composition as defined in claim 4 , wherein a protein transcribed from said protein-coding RNA is an enzyme.
13. The composition as defined in claim 1 , wherein said prokaryotic cells are bacterial cells.
14. The composition as defined in claim 1 , wherein said prokaryotic cells are Escherichia coli ( E. coli ).
15. The composition as defined in claim 1 , wherein said chemical agent is a transcription inducer for inducing RNA expression of said gene encoding at least the hairpin-like structure in said prokaryotic cells.
16. The composition as defined in claim 1 , wherein said chemical agent in said condition has a v/v concentration of 0.01% to 1%.
17. The composition as defined in claim 4 , wherein said non-coding RNA is useful for pharmaceutical or therapeutic application.
18. The composition as defined in claim 4 , wherein said protein-coding RNA is useful for pharmaceutical or therapeutic application.
19. The composition as defined in claim 4 , wherein said non-coding RNA is a precursor of microRNA.
20. The composition as defined in claim 19 , wherein said precursor of microRNA is a precursor of miR-302.
21. The composition as defined in claim 4 , wherein a protein transcribed from said protein-coding RNA is useful for pharmaceutical or therapeutic application.
22. The composition as defined in claim 17 , wherein said pharmaceutical or therapeutic application is cancer therapy and disease treatment.