Synonymous Modification Enables High Fidelity Expression of Biosensors and Probes with Repetitive Protein and Nucleotide Sequences

摘要: Repetitive nucleotide or amino acid sequences are often engineered into biosensors and probes to achieve functional readouts robust signal amplification. However, these repeated notoriously difficult construct maintain for stable expression. They easily deleted truncated, which occurs randomly is not a priori predictable, resulting in aberrant expression impacting the ability correctly detect interpret biological functions. Here, we introduce simple generally applicable approach solve this unappreciated problem by modifying of target mRNA make them non-repetitive but still (“synonymous”). We first demonstrate procedure designing cassette synonymous MS2 RNA aptamers, tandem coat proteins imaging show dramatic improvement reproducibility single detection live cells. The same extended enhance stability fluorescent containing FRET-pair proteins, upon great majority systems thus far field based. Using modified FRET biosensors, correct full-length sensors, eliminating truncation products that were assumed be due non-specific proteolytic cleavages. Importantly, interpretations sensor significantly different when correct, biosensor expressed. Thus method describe here should routinely employed generation sensors multiple, repetitive motifs state-of-the-art tools.