Difference between revisions of "Part:BBa K2916052"

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Toehold for Grapevine Endogenous Control 2.2

Toehold switches are synthetic riboregulators that regulate the expression of a downstream gene. They sequester the Ribosome Binding Site (RBS) and Start Codon by forming a secondary structure called a hairpin structure in its OFF state (acting as a “lock” for protein expression). A second strand of mRNA, complementary to the trigger binding region, needs to be present to activate the toehold. It will bind to that region and linearize the hairpin structure, freeing the RBS for the ribosomes to bind and allowing for translation of the downstream gene. Our toehold switches function at 37°C, and protein production is carried out in a cell-free PURE system. Reference Toehold switches: de-novo-designed regulators of gene expression, Green et al., 2014 In the encoding part, we have a toehold region, a hairpin region which contains a stem and a loop, a refolding region and a spacer. In design, it is crucial to exclude any ending codon in the refolding and spacer region. The refolding region allows ribosomes to access the protein coding region with less energy, thus increasing the kinetics of the reaction. Sequence and Features