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Abstract

A method for identifying a mutation-sensitive active region of a test protein, by providing a test nucleic acid construct comprising a regulatable promoter polynucleotide and a fusion polynucleotide comprising a test polynucleotide encoding the test protein fused to a reporter gene, wherein said fusion polynucleotide is operably associated with the promoter polynucleotide, wherein expression of the fusion polynucleotide in a selected host cell results in a specific phenotype and the presence of the reporter; mutagenizing the test nucleic acid construct to provide a mutagenized construct; transforming a selected host cell with the mutagenized construct to provide a transformed host cell; selecting a transformed host cell that exhibits the reporter, but which does not exhibit the specific phenotype; and sequencing a portion of the mutagenized construct from the selected transformed host cell to determine the alteration of the polynucleotide(s). The sequenced polynucleotides will exhibit alterations in one or more critical regions.

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First Claim