Depositing Lab: Paola Branduardi
The Easy-MISE toolkit, which stands for Easy-Modular Integrative fuSion-ready Expression toolkit, is a complete vector set leading to fast engineering procedures for the improvement of heterologous production in S. cerevisiae. It is characterized by a reduced design complexity thanks to a single-step assembly of the integration-ready expression cassette.
The final device obtained is a flexible double transcription unit cassette. Each transcription unit is transcribed in the opposite direction. Each integration cassette contains an upstream and a downstream homology sequence for targeted genome insertion. This is obtained by exploiting the CRISPR-Cas9 system and already available gRNA vectors from the EasyClone-MarkerFree Vector Set, targeting 11 previously well-characterized genome loci.
The toolkit allows an easy in-frame tag of every ORF of interest. This is achieved by the presence of a DNA spacer between each ORF and terminator that can be substituted with the desired sequence, for example with a GFP tag to study cell localization of the protein of interest, or with a specific ligand targeting the protein to a specific subcellular localization.
This kit will be sent as bacterial glycerol stocks in 96-well plate format.