Claims
- 1. A method of screening for an agent that modulates the activity of a target cell component, wherein the target cell component and a selectable marker are expressed by a recombinant cell, the method comprising co-encapsulating the agent in a micro-environment with the recombinant cell expressing the target cell component and detectable marker and detecting the effect of the agent on the activity of the cell component.
- 2. The method of claim 1, wherein the agent is an enzyme or small molecule.
- 3. The method of claim 1, wherein the agent is derived from a multispecific expression library.
- 4. The method of claim 2, wherein the enzyme is selected from the group consisting of lipases, esterases, proteases, glycosidases, glycosyl transferases, phosphatases, kinases, mono- and dioxygenases, haloperoxidases, lignin peroxidases, diarylpropane peroxidases, epozide hydrolases, nitrile hydratases, nitrilases, transaminases, amidases, and acylases.
- 5. The method of claim 1, wherein the agent inhibits the activity of the target cell component.
- 6. The method of claim 1, wherein the agent enhances the activity of the target cell component.
- 7. The method of claim 1, wherein the agent is expressed from a recombinant cell co-encapsulated with the recombinant cell expressing the target cell component and detectable marker.
- 8. The method of claim 7, wherein the recombinant cell is a eukaryotic cell.
- 9. The method of claim 7, wherein the recombinant cell is a prokaryotic cell.
- 10. The method of claim 1, wherein the micro-environment is a liposome, gel microdrop, bead, agarose, cell, ghost red blood cell or ghost macrophage.
- 11. The method of claim 10, wherein the liposomes are prepared from one or more phospholipids, glycolipids, steroids, alkyl phosphates or fatty acid esters.
- 12. The method of claim 11, wherein the phospholipids are selected from the group consisting of lecithin, sphingomyelin and dipalmitoyl.
- 13. The method of claim 11, wherein the steroids are selected from the group consisting of cholesterol, cholesterol and lanosterol.
- 14. The method of claim 1, wherein the detectable marker is a fluorescent dye, a visible dye, a bioluminescent material, a chemiluminescent material, a radioactive material, or an enzymatic substrate.
- 15. The method of claim 14, wherein the bioluminescent material is green fluorescent protein (GFP) or red fluorescent protein (RFP).
- 16. The method of claim 14, wherein detection of the fluorescent dye or a visible dye is carried out by fluorometric or spectrophotometric measurement.
- 17. The method of claim 1, wherein the protein is a transducing protein.
- 18. The method of claim 17, wherein the transducing protein is a G-protein.
- 19. The method of claim 1, wherein the recombinant cell is a eukaryotic cell.
- 20. The method of claim 1, wherein the recombinant cell is a prokaryotic cell.
- 21. A method for enriching for target DNA sequences containing at least a partial coding region for at least one specified activity in a DNA sample comprising:
a) co-encapsulating in a micro-environment a mixture of target DNA obtained from a mixture of organisms with a mixture of DNA probes comprising a detectable marker and at least a portion of a DNA sequence encoding at least one enzyme having a specified enzyme activity; b) incubating the co-encapsulated mixture under such conditions and for such time as to allow hybridization of complementary sequences; and c) screening for the specified activity.
- 22. The method of claim 21, further comprising transforming host cells with recovered target DNA to produce an expression library of a plurality of clones.
- 23. The method of claim 21, wherein the organisms are microorganisms.
- 24. The method of claim 23, wherein the microorganisms are uncultured microorganisms.
- 25. The method of claim 21, further comprising screening the expression library for the specified enzyme activity.
- 26. The method of claim 21, wherein the target DNA obtained from the DNA population is selected by:
a) converting double stranded DNA into single stranded DNA; b) recovering from the converted single stranded DNA, single stranded target DNA which hybridizes to probe DNA; c) converting recovered single stranded target DNA to double stranded DNA; and d) transforming a host cell with the double stranded DNA of c).
- 27. The method of claim 21, wherein screening is by FACS analysis.
- 28. The method of claim 21, wherein said target DNA is gene cluster DNA.
- 29. The method of claim 24, wherein the uncultured microorganisms are derived from an environmental sample.
- 30. The method of claim 23, wherein the uncultured microorganisms comprise a mixture of terrestrial microorganisms or marine microorganisms or airborne microorganisms, or a mixture of terrestrial microorganisms, marine microorganisms and airborne microorganisms.
- 31. The method of claim 22, wherein the clones comprise a construct selected from the group consisting of phage, plasmids, phagemids, cosmids, fosmids, viral vectors, and artificial chromosomes.
- 32. The method of claim 21, wherein the target DNA comprises one or more operons, or portions thereof, of the DNA population.
- 33. The method of claim 32, wherein the operon or portions thereof encodes a complete or partial metabolic pathway.
- 34. The method of claim 24, wherein the uncultured microorganisms comprise extremophiles.
- 35. The method of claim 34, wherein the extremophiles are selected from the group consisting of thermophiles, hyperthermophiles, psychrophiles, barophiles, and psychrotrophs.
- 36. The method of claim 26, wherein the host cell is selected from the group consisting of a bacterium, fungus, plant cell, insect cell and animal cell.
- 37. The method of claim 21, wherein the target DNA encodes a protein.
- 38. The method of claim 37, wherein the protein is an enzyme.
- 39. The method of claim 38 wherein the enzyme is selected from the group consisting of oxidoreductases, transferases, hydrolases, lyases, isomerases, and ligases.
- 40. The method of claim 21, wherein the micro-environment is a liposome, gel microdrop, bead, agarose, cell, ghost red blood cell or ghost macrophage.
- 41. The method of claim 40, wherein the liposomes are prepared from one or more phospholipids, glycolipids, steroids, alkyl phosphates or fatty acid esters.
- 42. The method of claim 41, wherein the phospholipids are selected from the group consisting of lecithin, sphingomyelin and dipalmitoyl.
- 43. The method of claim 41, wherein the steroids are selected from the group consisting of cholesterol, cholestanol and lanosterol.
- 44. The method of claim 21, wherein the detectable marker is a fluorescent dye, a visible dye, a bioluminescent material, a chemiluminescent material, a radioactive material, or an enzymatic substrate.
- 45. The method of claim 44, wherein the bioluminescent material is green fluorescent protein (GFP) or red fluorescent protein (RFP).
- 46. The method of claim 45, wherein detection of the fluorescent dye or a visible dye is carried out by fluorometric or spectrophotometric measurement.
- 47. A method of screening for an agent that modulates the interaction of a first test protein linked to a DNA binding moiety and a second test protein linked to a transcriptional activation moiety, comprising co-encapsulating the agent with the first test protein and second test protein in a suitable microenvironment and determining the ability of the agent to modulate the interaction of the first test protein linked to a DNA binding moiety with the second test protein covalently linked to a transcriptional activation moiety, wherein the agent enhances or inhibits the expression of a detectable protein, and wherein the enhancement or inhibition is detected by FACS analysis.
- 48. The method of claim 47, wherein the agent is an enzyme or small molecule.
- 49. The method of claim 48, wherein the enzyme is selected from the group consisting of lipases, esterases, proteases, glycosidases, glycosyl transferases, phosphatases, kinases, mono- and dioxygenases, haloperoxidases, lignin peroxidases, diarylpropane peroxidases, epozide hydrolases, nitrile hydratases, nitrilases, transaminases, amidases, and acylases.
- 50. The method of claim 47, wherein the agent inhibits the activity of the first protein or the second protein.
- 51. The method of claim 47, wherein the agent enhances the activity of the first protein or the second protein.
- 52. The method of claim 47, wherein the agent is expressed from a recombinant cell co-encapsulated with the recombinant cell expressing the target protein and detectable marker.
- 53. The method of claim 52, wherein the recombinant cell is a eukaryotic cell.
- 54. The method of claim 52, wherein the recombinant cell is a prokaryotic cell.
- 55. The method of claim 47, wherein the micro-environment is a liposome, gel microdrop, bead, agarose, cell, ghost red blood cell or ghost macrophage.
- 56. The method of claim 55, wherein the liposomes are prepared from one or more phospholipids, glycolipids, steroids, alkyl phosphates or fatty acid esters.
- 57. The method of claim 56, wherein the phospholipids are selected from the group consisting of lecithin, sphingomyelin and dipalmitoyl.
- 58. The method of claim 56, wherein the steroids are selected from the group consisting of cholesterol, chlorestanol and lanosterol.
- 59. The method of claim 47, wherein the detectable marker is a fluorescent dye, a visible dye, a bioluminescent material, a chemiluminescent material, a radioactive material, or an enzymatic substrate.
- 60. The method of claim 59, wherein the bioluminescent material is green fluorescent protein GFP) or red fluorescent protein (RFP).
- 61. The method of claim 59, wherein detection of the fluorescent dye or a visible dye is carried out by fluorometric or spectrophotometric measurement.
- 62. A method of identifying a molecule that modulates the activity of a target cell component comprising co-encapsulating the molecule and a cell expressing a target cell component of interest and a detectable marker, in a microenvironment under conditions that allow the molecule and the target cell component to interact, and detecting an effect of the molecule on the activity of the cell component, thereby identifying a molecule that modulates the activity of the target cell component.
Parent Case Info
[0001] This application is a continuation of U.S. patent application Ser. No. 09/098,206, filed Jun. 16, 1998, which is a continuation-in-part of U.S. patent application Ser. No. 08/876,276, filed Jun. 16, 1997, now abandoned.
Continuations (2)
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Number |
Date |
Country |
Parent |
09636778 |
Aug 2000 |
US |
Child |
10096701 |
Mar 2002 |
US |
Parent |
09098206 |
Jun 1998 |
US |
Child |
09636778 |
Aug 2000 |
US |
Continuation in Parts (1)
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Number |
Date |
Country |
Parent |
08876276 |
Jun 1997 |
US |
Child |
09098206 |
Jun 1998 |
US |