Internal ribosomal entry sites (IRES) are an important but poorly understood part of the eukaryotic translational machinery, allowing cap-independent translation initiation. Unfortunately, the Registry of Standard Biological Parts contains few IRESs and even those are poorly annotated. In this work, a group of IRESs were characterized for relative strength and submitted to the registry. To do this, we developed a methodology of determining relative IRES strength, as modeled by the methodology put forth by Kelly et al in the Journal of Biological Engineering (Kelly et al., 2001). Using flow cytometry, we determined the relative levels of fluorescent protein under the control of each IRES, as expressed in S. cerevisiae. Finally, we will show a practical example of how IRESs can be used in chemical engineering and synthetic biology. This work will not only be useful for other projects within our sponsor lab, but will also serve the greater synthetic biology community by initiating the growth of a library of IRESs and a protocol to allow contribution by all members of the community.
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