Yeast Golden Gate: Standardized Assembly of S. cerevisiae Transcriptional Units

Abstract

BBF RFC 88 describes a new standard for the assembly of basic Saccharomyces cerevisiae transcriptional units (TUs) consisting of a promoter/5’untranslated region (UTR), open reading frame (ORF), and 3’UTR/terminator. Note that we use the term “promoter” here to refer to both the promoter and the 5’ UTR, which we currently define as a single part. Future iterations of this standard will incorporate subdivision of currently defined parts e.g. into promoter and 5’ UTR. The standard makes use of the type IIS restriction enzyme BsaI to generate standardized and user­‐defined ‘signature overhangs’, thus enabling directional and seamless TU assembly. RFC88 is supported by the Yeast Standardized Collection of Parts for Expression (SCoPE), a repository of subcloned and sequence verified parts compatible with this assembly standard. The Yeast SCoPE is currently populated by a large number of S. cerevisiae promoters and terminators that facilitate expression and characterization of non­‐native ORFs.