Difference between revisions of "Team:Michigan/Design"

Revision as of 01:09, 19 September 2015

Design

While there are many genetic switches for detecting proteins, for example, the lac operon and tet regulators, to our knowledge, every genetic switch is unique to the protein it detects. Aptapaper seeks to create a switch that could have the same logic adapted to any protein, similar to how toehold switches can be adapted and respond to any sequence of trigger RNA. Toehold switches refer to an RNA switch that works by sequestering the ribosomal binding site and start codon in a hairpin, while leaving a portion of what the RNA trigger binds to exposed in a “toehold” region. This toehold region dramatically changes the kinematics of the system, making it much easier for the trigger to bind. After the initial binding, the hairpin is “unzipped” by the trigger¹. Exact design specs of the well optimized second generation toehold design are detailed in Figure 1. Because toehold switches can respond to any RNA trigger, Aptapaper attempted to use aptamers in conjunction with toehold switches to allow efficient and sensitive protein sensing. See different design approaches below.

Figure 1: Forward Engineered Switches Inspiring Aptapaper Switch Design

A.) RNA switch without trigger forms a stable hairpin structure, featuring a 15nt toehold region at 5’ end, 18nt total hairpin stem, 3nt AUG bubble (yellow), 15nt bubble sequestering 8nt ribosome binding site (green). Bubbles for AUG and ribosomal site allow switch to be activated by RNA triggers with any sequence. B.) RNA trigger (red) easily binds exposed toehold region. C.) RNA trigger binds to hairpin region of switch, forcing hairpin to unbind. D.) RNA trigger has fully bound switch, exposing the ribosome binding site so translation can be initiated1.

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