Patent ID: 7794971

Claim:
A method for cloning and stably maintaining a DNA sequence encoding a foreign product in a host cell, said method comprising: A. providing: (1) a host cell for conditional control of copy number of a recombinant plasmid that is introduced into said host cell, wherein said host cell comprises a poly(A) polymerase gene that (a) is operably joined to a conditionally inducible promoter, (b) is located on a chromosome of said host cell, and (c) catalyzes polyadenylation of an antisense RNA molecule that affects initiation of replication from an origin of replication (ori) in said recombinant plasmid wherein the copy number of the recombinant plasmid in the host cell is lower when the poly(A) polymerase gene is uninduced than when the poly(A) polymerase gene is induced, wherein said host cell additionally comprises an RNA polymerase gene operably joined to a conditionally-inducible promoter that is different from the conditionally-inducible promoter to which the poly(A) polymerase gene is operably joined; and (2) a DNA sequence encoding a foreign product; B. ligating said DNA sequence encoding the foreign product into a plasmid, which DNA sequence is operably joined to an RNA polymerase promoter recognized by the RNA polymerase, so as to obtain a recombinant plasmid, wherein replication level and copy number of said recombinant plasmid in the host cell is regulated by an antisense RNA that is polyadenylated by a poly(A) polymerase encoded by the poly(A) polymerase gene that is operably joined to the conditionally-inducible promoter in said host cell; C. introducing said recombinant plasmid into said host cell; and D. growing said host cell containing said recombinant plasmid that contains said DNA sequence under conditions wherein said conditionally inducible promoter that is operably joined to said poly(A) polymerase gene in said host cell is not induced, wherein the host cell is E. coli , the antisense RNA molecule is copA or RNAI and the recombinant plasmid is a R1-type or colE1-type plasmid, thereby maintaining said recombinant plasmid at low copy in said host cell under said conditions.