DNA fragments containing gene having function relating to autonomous proliferation of plasmid
View Patent ↗Provided are a DNA fragment containing a gene relating the autonomous replication of a plasmid, and a DNA fragment containing a gene relating to the autonomous replication of a plasmid wherein a mutation point capable of increasing the number of copies of a plasmid is present in at least one site in the gene. Also provided are a DNA fragment containing a gene having a function relating to the autonomous replication of a plasmid within the bacteria of the genus Rhodococcus , a DNA fragment wherein a mutation point capable of increasing the number of copies of a plasmid is present in at least one site in the gene, and a plasmid carrying a DNA fragment wherein a mutation point capable of increasing the number of copies of a plasmid is present in at least one site of the gene.
1. An isolated polynucleotide which is at least 95% homologous to SEQ ID NO: 7 or which hybridizes to the full complement of SEQ ID NO: 7 under stringent conditions, wherein stringent conditions comprise washing in 0.1×SSC and 0.1% SDS at 60° C.;
wherein said isolated polynucleotide contains a point mutation of the polynucleotide sequence of SEQ ID NO: 7 and the point mutation changes the 71 st Gly of the amino acid sequence of SEQ ID NO: 8 to Ser, and
wherein the isolated polynucleotide increases plasmid copy number when incorporated into plasmid in Rhodococcus when compared to the plasmid copy number in Rhodococcus containing a plasmid having the unaltered polynucleotide sequence of SEQ ID NO: 7.
2. The isolated polynucleotide of claim 1 , which comprises a point mutation at nucleotide 211 of SEQ ID NO: 7 so that nucleotide 211 is thymine.
3. A vector comprising the isolated polynucleotide of claim 1 or 2 .
4. An isolated host cell comprising the vector of claim 3 .
5. The isolated host cell of claim 4 which is Rhodococcus.
6. A method for increasing plasmid copy number in Rhodococcus comprising transforming a Rhodococcus cell with the vector of claim 3 .
7. An isolated polynucleotide which comprises SEQ ID NO: 2.
8. An isolated polynucleotide which comprises SEQ ID NO: 9.
9. A vector comprising the isolated polynucleotide of claim 7 or 8 .
10. An isolated host cell comprising the vector of claim 9 .
11. The isolated host cell of claim 10 which is Rhodococcus.
12. A method for increasing plasmid copy number in Rhodococcus comprising transforming a Rhodococcus cell with the vector of claim 9 .
13. Plasmid pLK006 identified by accession number FERM BP-8085.