Host-vector system antibiotic-free ColE1 plasmid propagation

摘要

A host-vector system that uses the RNA-based copy number control mechanism of ColE1-type plasmids for regulating the expression of a marker gene allows for antibiotic-free selection of plasmids and is useful for production of plasmid DNA and recombinant proteins.

主权项

1. A non-naturally occurring bacterial cell containing
i) a DNA sequence encoding a marker protein, the expression of which is to be regulated, operably associated to ii) a DNA sequence encoding a part of an RNA II sequence
a) that is complementary to an RNA I sequence from a plasmid with a ColE1 origin of replication, andb) that is positioned upstream or downstream of the ribosomal binding site of the DNA sequence of i), such that in the absence of RNA I in said bacterial cell the mRNA of the DNA sequence of i) is translated into a marker protein, wherein said part is a two loop structure of the RNA II sequence, wherein said bacterial cell contains said DNA sequences i) and ii) integrated in its genome.