A Yeast Logic Toolkit for Fast and Efficient Assembly of Genetic Circuits

Synthetic biologists aim for standardization and simplicity in order to ensure reproducibility and reliability. Toolkits offer both, making it easier to share DNA sequences between labs while minimizing context dependent effects from sequences surrounding the construct of interest. The Yeast Toolkit (YTK) for the model organism Saccharomyces cerevisiae has found wide adoption within the scientific community. While the Toolkit and its existing extensions offer a large variety of different parts, there is a lack of characterized logic gates and insulator containing backbones for the assembly of genetic circuits with the yeast toolkit syntax. In this work, we present the Yeast Logic Toolkit (YLT). YLT contains 40 pre-assembled backbones with dropouts, 109 parts for NOT/NOR gate assembly, and a subdivided promoter Type 2 part that allows more flexible 5'UTR design. For insulation, the backbones contain spacer sequences and self-cleaving ribozymes which buffer against transcriptional read-through from upstream sequences. Due to our subdivision of Type 2 parts, we are able to manipulate the 5' untranslated region without a cloning scar upstream of the start codon. We also characterized the NOT and NOR gates from the Cello library for yeast in the sequence context of the yeast toolkit cloning scheme. This toolkit extension will make it easier and faster for scientists to assemble genetic circuits in yeast while providing a minimum of context dependent effects such as genetic read-trough due to the standardization of up- and downstream sequences surrounding the transcriptional units.