CH391L/S12/In vitro Selection

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Once diversity is created, the selection must must allow function variants to become a larger percentage of the pool.  This step is often the most difficult to design.   
Once diversity is created, the selection must must allow function variants to become a larger percentage of the pool.  This step is often the most difficult to design.   
=====Affinity=====
=====Affinity=====
-
The easiest function to enrich for is affinity for a ligand.  To select for binders (aptamers for nucleic acids, antibodies and others for proteins)one exposes the pool of potential binders to a fixed ligand.  The best binders affix to the ligand while weaker binders are washed away.  Those that remain can be amplified for further round of selection.  For nucleic acids, the binder itself can be subject to amplification, as it is both the information-carrying and function-carrying molecule.  For protein binders, the scheme must include some sort of linking of the information-carrying nucleic acid and the function-carrying protein.  Come examples of this linking are phage-display, cell-surface-display, and ribosome-display.
+
The easiest function to enrich for is affinity for a ligand.  To select for binders (aptamers for nucleic acids; antibodies and others for proteins,) one exposes the pool of potential binders to a fixed ligand.  The best binders affix to the ligand while weaker binders are washed away.  Those that remain can be amplified for further round of selection.  For nucleic acids, the binder itself can be subject to amplification, as it is both the information-carrying and function-carrying molecule.  For protein binders, the scheme must include linking of the information-carrying nucleic acid to the function-carrying protein.  Some examples of this linking are phage display, cell-surface display, and ribosome display.
=====Protection=====
=====Protection=====
=====Selective Amplification=====
=====Selective Amplification=====

Revision as of 21:03, 28 January 2012

Contents

Overview of in vitro selection

Library Generation

Randomized Oligodeoxynucleotides
Mutagenic PCR
Gene Shuffling
Neutral Drift

Increased Representation

Once diversity is created, the selection must must allow function variants to become a larger percentage of the pool. This step is often the most difficult to design.

Affinity

The easiest function to enrich for is affinity for a ligand. To select for binders (aptamers for nucleic acids; antibodies and others for proteins,) one exposes the pool of potential binders to a fixed ligand. The best binders affix to the ligand while weaker binders are washed away. Those that remain can be amplified for further round of selection. For nucleic acids, the binder itself can be subject to amplification, as it is both the information-carrying and function-carrying molecule. For protein binders, the scheme must include linking of the information-carrying nucleic acid to the function-carrying protein. Some examples of this linking are phage display, cell-surface display, and ribosome display.

Protection
Selective Amplification
FACS

Confinement of Function

Binding
Cis-action
Cellularization

===In vitro Compartmentalization=======History

Nucleic Acids

Aptamers

Ellington 1990

Ribozymes

Bartel 1993

Proteins

Antibodies

Accepting suggestions

DNA Modifying Proteins

Tawfik 1998

Other Enzymatic Functions

Griffiths 2003 Griffiths 2003 Schema

Self-Replication

Ghadessy 2001


References

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