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Promoter analysis by saturation mutagenesis

Abstract

Gene expression and regulation are mediated by DNA sequences, in most instances, directly upstream to the coding sequences by recruiting transcription factors, regulators, and a RNA polymerase in a spatially defined fashion. Few nucleotides within a promoter make contact with the bound proteins. The minimal set of nucleotides that can recruit a protein factor is called a cis-acting element. This article addresses a powerful mutagenesis strategy that can be employed to define cis-acting elements at a molecular level. Technical details including primer design, saturation mutagenesis, construction of promoter libraries, phenotypic analysis, data analysis, and interpretation are discussed.

Abbreviations

bop :

bacterioopsin

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Correspondence to Nitin S. Baliga.

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Published: December 22, 2001

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Baliga, N.S. Promoter analysis by saturation mutagenesis. Biol Proced Online 3, 64–69 (2001). https://doi.org/10.1251/bpo24

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Indexing terms

  • promoter regions (genetics)
  • mutagenesis
  • polymerase chain reaction