Compositions and methods for the identification and selection of nucleic acids and polypeptides

A method for evolving a desired protein sequence comprising:

• providing at least two candidate mRNA molecules, wherein at least one of said mRNA molecules contains at least one codon selected from the group consisting of a stop codon and a pseudo stop codon;
• translating at least two of said candidate mRNA molecules to generate at least one translated protein;
• linking at least one of said candidate mRNA molecules to its corresponding translated protein via a tRNA molecule to form at least one cognate pair, wherein at least one of said candidate mRNA molecules is connected to said tRNA molecule by a crosslinker;
• identifying one or more of said cognate pairs based upon the properties of said translated protein or said mRNA molecule;
• identifying a molecule selected from the group consisting of one or more of the following:
• an mRNA molecule of said selected cognate pair, a nucleic acid molecule complementary to said mRNA molecule and a nucleic acid molecule homologous to said mRNA molecule, thereby identifying said desired protein or said desired nucleic acid molecule;
• providing a plurality of cognate pairs,
• binding at least of said plurality of cognate pairs with one or more binding agents;
• selecting said desired or protein nucleic acid molecule based upon a reaction or lack of a reaction to said one or more binding agents, thereby selecting a first desired cognate pair;
• recovering said first desired cognate pair to generate a recovered cognate pair;
• amplifying a first nucleic acid component of said recovered cognate pair;
• producing a second nucleic acid component, wherein said second nucleic acid component comprises said first nucleic acid component with one or more variations;
• producing a second protein by translating said second nucleic acid component;
• linking said second protein with said second nucleic acid component to generate a second desired cognate pair;
• and
• obtaining the desired protein sequence by re-selecting said second desired cognate pair based upon at least one desired property.

A method of forming a psoralen monoadduct on a nucleic acid, comprising:

• providing a first nucleic acid and a second nucleic acid,
• wherein said first nucleic acid and said second nucleic acid are substantially complementary to each other,
• wherein said first nucleic acid comprises one or more uridine monoadduct targets, and
• wherein said second nucleic acid comprises at least one pseudoridine
• hybridizing said first nucleic acid and said second nucleic acid in the presence of psoralen to form a hybrid;
• irradiating said hybrid with ultraviolet light, thereby forming said psoralen monoadduct on said first nucleic acid.

A method of producing a psoralen monoadduct or a crosslink, comprising:

• providing a first nucleic acid and a second nucleic acid;
• wherein said first nucleic acid and said second nucleic acid are substantially complementary to each other;
• wherein said first nucleic acid comprises one or more uridine monadduct targets or crosslink targets and one or more uridine monoadduct non-targets or crosslink non-targets;
• wherein said uridine monoadduct non-targets or crosslink non-targets are operable to be replaced with one or more pseudouridines;
• replacing one or more of said uridine monoadduct non-targets or crosslink non-targets with pseudouridine;
• hybridizing said first nucleic acid and said second nucleic acid in the presence of psoralen- to form a hybrid;
• irradiating said hybrid, thereby forming said psoralen monoadduct or said crosslink on said first nucleic acid on said targets, while protecting said nontargets.