TIS_5

This Biomodule encodes for a yeast transcription initiation site (TIS) of the MIAMI Project collection. This TIS was characterized in vivo in S.cerevisiae [1] using the protocol described in [2]. Briefly, the TIS was cloned downstream the pPGK1 promoter in the MIAMI – Standard reporter gene cassette [3] and integrated into S.cerevisiae genome by EasyClone marker-free integration as described in [4]. TIS’s strength was assessed in vivo over 24 hours and results re-scaled using a fluorescein standard curve as described in [5].

MIAMI collection was designed by members of the MIAMI consortium supported by the EU Horizon 2020 research and innovation programme. Grant agreement No 814645.

Reference:
[1] S.cerevisiae CEN.PK2-1C
[2] MIAMI's Standard Operating Procedures
[3] MIAMI - Standard reporter gene cassette
[4] Mathew M Jessop‐Fabre et al. EasyClone‐MarkerFree: A vector toolkit for marker‐less integration of genes into Saccharomyces cerevisiae via CRISPR‐Cas9. Biotechnol J. 2016 Aug; 11(8): 1110–1117.Link
[5] Whole cell fluorescence reading Ex485nm/Em530nm with plate reader