Bacterial strains for DNA production

Abstract

[Problem] A composition for the production of plasmid nucleic acids, and a method for preparing and using the same, is provided. [Solution] Engineered bacterial strains and vectors for enhanced plasmid DNA production are provided herein. [Selected Figure] None

Claims

1. A genetically modified microorganism comprising a nucleic acid containing a prsA variant, wherein the nucleic acid encodes a protein having at least 70% sequence identity with prsA* (SEQ ID NO: 23), and / or the nucleic acid encodes a protein having at least 95% sequence identity with prsA* (SEQ ID NO: 24).

2. The genetically modified microorganism according to Claim 1, wherein the microorganism has a genome in which the repressor gene purR is disrupted or deleted.

3. The genetically modified microorganism according to claim 2, wherein the purR comprises the sequence of SEQ ID NO: 25, and / or the prsA variant has 90% sequence identity with respect to prsA* (SEQ ID NO: 23).

4. A genetically modified microorganism according to any one of claims 1 to 3, wherein the EcoKI restriction system, endA and / or recA are deleted from the genome.

5. The genetically modified microorganism according to any one of claims 1 to 3, wherein the genetically modified microorganism is a recombinant strain of Escherichia coli (E. coli).

6. A genetically modified microorganism containing the prsA variant, wherein the microorganism has a genome in which the repressor gene purR is disrupted.

7. A recombinant strain of Escherichia coli (E. coli) comprising an E. coli genome having at least the following gene deletions: endA (ΔendA) and recA (ΔrecA), wherein the E. coli optionally contains the prsA variant.

8. The recombinant strain according to claim 7, wherein the E. coli is derived from MG1655, and the E. coli genome optionally includes a nucleic acid sequence having at least 80% identity with the MG1655 genome.

9. The recombinant strain according to claim 7 or 8, wherein the EcoKI restriction system is deleted from the genome of E. coli, and / or the purR sequence is deleted from the genome of E. coli, and optionally the E. coli genome has the nucleic acid sequence of Sequence ID No. 25 deleted with respect to the MG1655 genome.

10. The recombinant strain according to claim 7, wherein the E. coli genome comprises the nucleic acid sequence of SEQ ID NO:

23.

11. The recombinant strain according to any one of claims 7 to 10, wherein the E. coli genome further comprises at least one gene deletion selected from the group including mrr, hsdR, hsdM, hsdS, symE, and mcrBC.

12. The recombinant strain according to claim 7, wherein the recombinant strain comprises a plasmid having the genotype: |<repA101|ori101_ts|<recA|<bla|<tetR|<P(tetR)|P(tet)>|gamma>|beta>|exo>|60a>|.

13. A recombinant plasmid comprising the genotype: |<repA101|ori101_ts|<recA|<bla|<tetR|<P(tetR)|P(tet)>|gamma>|beta>|exo>|60a>|.

14. A nucleic acid containing a prsA variant, wherein the nucleic acid has 70% to 99% sequence identity with respect to prsA* (SEQ ID NO: 23).

15. The nucleic acid according to claim 14, wherein the nucleic acid encodes a protein having at least 95% sequence identity with respect to prsA* (SEQ ID NO: 24).

16. The nucleic acid according to claim 14, wherein the nucleic acid has at least 80%, 90%, or 95% sequence identity with respect to prsA* (sequence number 23).

17. The nucleic acid according to claim 14, wherein the nucleic acid encodes a protein having 100% sequence identity with SEQ ID NO: 23 or 100% sequence identity with SEQ ID NO:

24.

18. A genetically modified microorganism, recombinant strain, or nucleic acid according to any one of claims 1 to 17, further comprising an open reading frame (ORF) encoding the target mRNA.

19. A manipulated nucleic acid vector containing a nucleic acid sequence having at least 95% sequence identity with sequence number 10.

20. An engineered nucleic acid vector containing a nucleic acid sequence having at least 95% sequence identity with sequence number 11.

21. A manipulated nucleic acid vector containing the nucleic acid sequence of sequence number 10.

22. A manipulated nucleic acid vector containing the nucleic acid sequence of sequence number 11.