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 useful for regulating eukaryotic promoter-driven gene expression in prokaryotes, comprising:
a chemical agent, containing a structure similar to 3-morpholinopropane-1-sulfonic acid (MOPS), ethanol, glycerin, or a mixture thereof.
2 . The composition as defined in claim 1 , wherein said chemical agent is a transcription inducer for gene expression.
3 . The composition as defined in claim 1 , wherein said composition further comprises a bacterial culturing medium.
4 . The composition 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 composition as defined in claim 3 , wherein said bacterial culturing medium is Luria-Bertani (LB) broth.
6 . The composition as defined in claim 1 , wherein said gene encodes at least a protein or non-coding RNA, or both, which is useful for pharmaceutical or therapeutic application.
7 . The composition as defined in claim 6 , wherein said non-coding RNA is useful for pharmaceutical or therapeutic application.
8 . The composition as defined in claim 6 , wherein said non-coding RNA is a miR-302 homologue.
9 . The composition as defined in claim 6 , wherein said protein or peptide is useful for pharmaceutical or therapeutic application.
10 . The composition as defined in claim 6 , wherein said pharmaceutical or therapeutic application is selected from generating pluripotent stem cells, stem cell research and therapy, cancer therapy and disease treatment, wound healing treatment, or generating high-yield food and drug supplies.
11 . A composition for regulating eukaryotic promoter-driven gene expression in prokaryotes, comprising:
(a) at least a chemical agent capable of inducing or enhancing eukaryotic promoter-driven gene expression, wherein said chemical agent containing a structure similar to 3-morpholinopropane-1-sulfonic acid (MOPS), ethanol, glycerin, or a mixture thereof; and
(b) a plurality of prokaryotic cells, said plurality of prokaryotic cells containing at least a gene mediated by a eukaryotic promoter-driven expression mechanism;
wherein (a) and (b) are mixed together under a condition to induce the eukaryotic promoter-driven gene expression of said gene in said prokaryotic cells.
12 . The composition as defined in claim 11 , wherein each one of said prokaryotic cells has at least an inducible gene expression composition.
13 . The composition as defined in claim 12 , wherein said inducible gene expression composition is a vector containing a eukaryotic promoter.
14 . The composition as defined in claim 11 , wherein said gene encodes at least a non-coding RNA or protein-coding RNA, or both.
15 . The composition as defined in claim 14 , wherein said non-coding RNA contains at least a sequence containing 30% to 100% homology to a microRNA.
16 . The composition as defined in claim 14 , wherein said non-coding RNA is a small hairpin RNA.
17 . The composition as defined in claim 14 , wherein said protein-coding RNA contains at least a sequence containing 30% to 100% homology to a microRNA.
18 . The composition as defined in claim 14 , wherein said protein-coding RNA contains 30% to 100% homology to a cDNA sequence of a eukaryotic gene.
19 . The composition as defined in claim 14 , wherein said non-coding RNA or protein-coding RNA is isolatable from said prokaryotes.
20 . The composition as defined in claim 11 , wherein said gene encodes at least a protein or peptide, or both.
21 . The composition as defined in claim 20 , wherein said protein contains at least a peptide sequence.
22 . The composition as defined in claim 20 , wherein said protein or peptide is isolatable from said prokaryotes.
23 . The composition as defined in claim 20 , wherein said protein is an enzyme or an antibody.
24 . The composition as defined in claim 20 , wherein said protein contains a peptide sequence similar to insulin.
25 . The composition as defined in claim 20 , wherein said protein contains a peptide sequence similar to a growth factor.
26 . The composition as defined in claim 11 , wherein said prokaryotic cells are bacterial cells.
27 . The composition as defined in claim 11 , wherein said prokaryotic cells are Escherichia coli ( E. coli ).
28 . The composition as defined in claim 11 , wherein said chemical agent is a transcription inducer for gene expression.
29 . The composition as defined in claim 11 , wherein said chemical agent contains a structure similar to 3-morpholinopropane-1-sulfonic acid (MOPS), ethanol or glycerin, or a mixture thereof.
30 . The composition as defined in claim 11 , wherein said eukaryotic promoter is a type II RNA polymerase (pol-2) equivalent or a pol-2 compatible viral promoter.
31 . The composition as defined in claim 30 , wherein said pol-2 compatible viral promoter is a cytomegaloviral (CMV) promoter or a retroviral long terminal repeat (LTR) promoter.
32 . The composition as defined in claim 11 , wherein said eukaryotic promoter-driven expression is a cellular mechanism including RNA transcription and protein translation.
33 . The composition as defined in claim 11 , wherein said condition is a bacterial culturing condition in Luria-Bertani (LB) broth at 37° C.
34 . The composition as defined in claim 11 , wherein said chemical agent in said condition has a v/v concentration of 0.01% to 1%.
35 . The composition as defined in claim 14 , wherein said non-coding RNA is useful for pharmaceutical or therapeutic application.
36 . The composition as defined in claim 14 , wherein said protein-coding RNA is useful for pharmaceutical or therapeutic application.
37 . The composition as defined in claim 14 , wherein said non-coding RNA is a miR-302 homologue.
38 . The composition as defined in claim 15 , wherein said microRNA is a miR-302 homologue.
39 . The composition as defined in claim 17 , wherein said microRNA is a miR-302 homologue.
40 . The composition as defined in claim 20 , wherein said protein or peptide is useful for pharmaceutical or therapeutic application.
41 . The composition as defined in claim 35 , wherein said pharmaceutical or therapeutic application is selected from generating pluripotent stem cells, stem cell research and therapy, cancer therapy and disease treatment, wound healing treatment, or generating high-yield food and drug supplies.
42 . The composition as defined in claim 36 , wherein said pharmaceutical or therapeutic application is selected from generating pluripotent stem cells, stem cell research and therapy, cancer therapy and disease treatment, wound healing treatment, or generating high-yield food and drug supplies.
43 . The composition as defined in claim 40 , wherein said pharmaceutical or therapeutic application is selected from generating pluripotent stem cells, stem cell research and therapy, cancer therapy and disease treatment, wound healing treatment, or generating high-yield food and drug supplies.