Genetically encoded biosensors for detection of polyketides
The present disclosure relates to high-throughput detection of polyketides using genetically encoded biosensors.
1. A biosensor system comprising:
a nucleic acid encoding a genetically modified MphR gene sequence, wherein the nucleic acid comprises at least one genetic mutation comprising at least one nucleotide change in the ribosome binding site sequence when compared to a wild-type MphR gene sequence; and
a reporter gene whose transcription is under the control of a promoter region which is regulated by a wild-type MphR transcription factor.
2. The biosensor system of claim 1 , wherein the nucleic acid encoding the genetically modified MphR gene sequence and the reporter gene are located on one recombinant DNA vector.
3. The biosensor system of claim 1 , wherein the reporter gene is a gene coding for chloramphenicol acetyltransferase, beta-galactosidase, luciferase or green fluorescent protein (GFP).
4. The biosensor system of claim 3 , wherein the reporter gene is a gene coding for green fluorescent protein (GFP).
5. The biosensor system of claim 1 , wherein the at least one nucleotide change in the ribosome binding site sequence is selected from A1G, A1T, A1C, G2T, G2A, A3C, A3G, A4T, G5T, G6T, or a combination thereof.
6. The biosensor system of claim 1 , wherein the at least one nucleotide change in the ribosome binding site sequence is selected from A1G, A4T, or a combination thereof.
7. A genetically modified host cell comprising:
a nucleic acid encoding the biosensor system of claim 1 .
8. The cell of claim 7 , wherein the at least one nucleotide change in the ribosome binding site sequence is selected from A1G, A1T, A1C, G2T, G2A, A3C, A3G, A4T, G5T, G6T, or a combination thereof.
9. The cell of claim 7 , wherein the cell is E. coli.
10. The cell of claim 7 , wherein the cell is Streptomyces.
11. A method for detecting a polyketide, comprising:
introducing into a cell:
a nucleic acid encoding a genetically modified MphR gene sequence, wherein the nucleic acid comprises at least one nucleotide change in the ribosome binding site sequence when compared to a wild-type MphR gene sequence;
and
a reporter gene whose transcription is under the control of a promoter region which is regulated by a wild-type MphR transcription factor; and
detecting the presence or absence of the polyketide based on the differential expression of the reporter gene in comparison to a cell comprising the wild-type MphR gene sequence.
12. The method of claim 11 , wherein the at least one nucleotide change in the ribosome binding site sequence is selected from A1G, A1T, A1C, G2T, G2A, A3C, A3G, A4T, G5T, G6T, or a combination thereof.
13. The method of claim 11 , wherein the cell is E. coli.
14. The method of claim 11 , wherein the cell is Streptomyces.