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Screening methods in eucaryotic cells

  • US 5,989,814 A
  • Filed: 04/01/1997
  • Issued: 11/23/1999
  • Est. Priority Date: 04/01/1997
  • Status: Expired due to Term
First Claim
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1. A method of screening a library of nucleic acid fragments in eucaryotic cells, comprising:

  • (1) transforming the library of nucleic acid fragments into primary cells, wherein the cells are procaryotic or fungi;

    (2) culturing the primary cells under conditions whereby the copy number of nucleic acid fragments is amplified to an average of at least 200 copies per transformed cell;

    (3) contacting the transformed primary cells with a population of eucaryotic cells, which lack capacity for episomal replication of the transferred nucleic acid fragments, under conditions whereby outersurfaces of the transformed primary cells and eucaryotic cells fuse and contents of the transformed primary cells including at least some of the library of nucleic acid fragments are transferred to the eucaryotic cells;

    (4) screening the nucleic acid fragments in the eucaryotic cells to isolate one or more eucaryotic cells having a desired property conferred by one or more members of the library of nucleic acid fragments, an expression product thereof, or a secondary metabolite of an expression product;

    (5) lysing the one or more eucaryotic cells to release the one or more members of the library of nucleic acid fragments and electroporating the nucleic acid fragments into further procaryotic cells;

    (6) propagating the further procaryotic cells to amplify the one or more members of the library of nucleic acid fragments, which confer the desired property.

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