Generation of a Library of Mutants - Doping DNA Template with Non-wild Type Nucleotides
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Synthesis of RNA
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Protein Binding Reaction and Separation of Bound RNA
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Analysis of Iodine-cleaved Phosphorothioate Products for Detection of Mutant Nucleotide Positions
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Results: Characterization of Protein Binding Site in RNA
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Conclusion
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The overall goal of this phosphorothioate approach in RNA is to perform saturation mutagenesis in a single step. This method can help answer key questions in the molecular biology field such as gene regulation. The main benefit of this technique i
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Proteins that bind specific RNA sequences play critical roles in gene expression. Detailed characterization of these binding sites is crucial for our understanding of gene regulation. Here, a single-step approach for saturation mutagenesis of protein-binding sites in RNA is described. This approach is relevant for all protein-binding sites in RNA.