Part:BBa_K319005

yeast mid-length ADH1 promoter

This is the complete ADH1 promoter including the distal and proximal portions and their regulatory elements.

Although there are other versions of the ADH1 promoter already available in the registry, they are longer than needed for optimal expression from the promoter. Furthermore, since these promoters are longer, they are inhibited by ethanol which is used as a source of energy during the ethanol phase of the yeast life cycle. This information has been published by Ruohonen et al. The length that was chosen for this ADH1 promoter is more manageable for the construction of larger networks, and is not inhibited by ethanol. We characterized the expression of the ADH1 promoter by cloning it in front of yEGFP with a ADH1 terminator. This construct was cloned into a pRS403 vector and integrated into the YPH500 genome. The resulting strain was innoculated in 3 mL of YPD + 2% Glucose + 2% Adenine overnight. The overnight culture was reinoculated into 2 mL of the same media at an OD of 0.02. After three hours the strain was analyzed on a Cyan ADP flow cytometer, 488 laser was used for exitation and emission was captured with a standard FITCH filterset. We observed roughly a 100 fold increase in GFP expression over wild type YPH500.

Figure 1: Fluorescence of wild type YPH500

Figure 2: Fluorescence of ADH1 promoter driving yEGFP expression in a YPH500 background.

References

Ruohonen et al. Modifications to the ADH1 promoter of Saccharomyces cerevisiae for efficient production of heterologous proteins. Journal of Biotechnology (1995) vol. 39 (3) pp. 193-203

Usage and Biology

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 93