CA 1195626

This patent was assigned to Novartis

CA 1195626

Actual granted independent claims
Claim 1
A recombinant DNA cloning vector comprising: a) a eukaryotic promoter;
b) one or two different structural genes and associated control sequence that convey resistance to either or both antibiotics hygromycin B and G418 when transformed into a host cell that is sensitive to either or both antibiotics for which resistance is conveyed, said host cell being susceptible to transformation, cell division, and culture; and
c) a prokaryotic replicon, said replicon being functional when said host cell is prokaryotic, subjected to the limitations

  • that the one or two structural genes and associated control sequence are adjacent to and, in a eukaryotic host cell, transcribed from the eukaryotic promoter,
  • that a single gene and associated control sequence conveys resistance to either but not both hygromycin B and G418, and
  • that the gene conveying resistance to G418 does not code for the enzyme phosphotransferase.
Claim 16A transformed host cell which comprises a recombinant DNA cloning vector comprising: a) a eukaryotic promoter;
b) one or two different structural genes and associated control sequence that convey resistance to either or both antibiotics hygromycin B and G418 when transformed into a host cell that is sensitive to either or both antibiotics for which resistance is conveyed, said host cell being susceptible to transformation, cell division, and culture; and
c) a prokaryotic replicon, said replicon being functional when said host cell is prokaryotic, subjected to the limitations

  • that the one or two structural genes and associated control sequence are adjacent to and, in a eukaryotic host cell, transcribed from the eukaryotic promoter,
  • that a single gene and associated control sequence conveys resistance to either but not both hygromycin B and G418, and
  • that the gene conveying resistance to G418 does not code for the enzyme phosphotransferase.
Claim 33A restriction fragment selected from the group consisting of:
a) the 7.5 kb Bgl II restriction fragment of plasmid pKC203;
b) the 2.75 kb Bgl II/Sal I restriction fragment of plasmid pKC203;
c) the 1.51 kb Sac I/Bgl II restriction fragment of plasmid pKC222; and
d) the 1.65 kb EcoR I/Sal I restriction fragment of plasmid pKC222.
Claim 34A plasmid selected from the group consisting of plasmid pKC203, pKC222, pKC257, pKC259, pKC261, pKC264, pKC275, and pSC70111 . 11 The plasmids pSC701 contains the 7.5 Bgl II fragment with resistance genes for both hygromycin and G418. The plasmid pKC259 also confers resistance to both antibiotics. The remaining plasmids all convey resistance to hygromycin B only. From them, plasmids pKC257, pKC259, pKC261 are functional in prokaryotes, and plasmids pKC264 and pKC275 are functional in eukaryotic host cells.
Claim 35A method for producing post translationally modified polypeptide, which comprises: a) transforming a eukaryotic cell with a recombinant DNA cloning vector comprising:

  1. a eukaryotic promoter;
  2. one or two different structural genes and associated control sequence that convey resistance to either or both antibiotics hygromycin B and G418 when transformed into a host cell that is sensitive to either or both antibiotics for which resistance is conveyed, said host cell being susceptible to transformation, cell division, and culture; and
  3. a prokaryotic replicon, said replicon being functional when said host cell is prokaryotic,

subjected to the limitations

  • that the one or two structural genes and associated control sequence are adjacent to and, in a eukaryotic host cell, transcribed from the eukaryotic promoter,
  • that a single gene and associated control sequence conveys resistance to either but not both hygromycin B and G418, and
  • that the gene conveying resistance to G418 does not code for the enzyme phosphotransferase, and

b) culturing the transformed cell.