Part:BBa_K2765026

GAL1p+yno1+CYC1t

This part consists of the promoter gal1, redox sensor yno1 and the terminator cyc1. The promoter gal1 is an constitutive promoter from Saccharomyces cerevisiae CENPK2-1C.yno1（yeast NADPH oxidase 1） is located in the endoplasmic reticulum (ER) and produces superoxide in a NADPH-dependent fashion encodes a genuine NADPH oxidase (NOX enzymes),(see the details in part:BBA-K2765007).And thus the target gene yno1 is induced by galactose and repressed by glucose.as shown in Fig.1.

We cloned yno1 from the genome of CEN.PK2-1C,then we ligated the gene yno1 with pESC plasmid,and cloned Pgal1-yno1-cyc1t from pESC plasmid.as shown in Fig.2.

After sequencing verification, we transferred the correct plasmid into yeast for expression. Then we used the galactose induction medium to induce the Gal1 promoter to overexpress the target gene, which increased the ROS content in the Saccharomyces cerevisiae cell.We used a suitable concentration of galactose to induce overexpression of the target gene, resulting in an increase in ROS accumulation compared to the control group.as shown in Fig.3.

References:

[1]Mark Rinnerthalera, Yno1p/Aim14p, a NADPH-oxidase ortholog, controls extramitochondrial reactive oxygen species generation, apoptosis, and actin cable formation in yeast ，2012

[2]Zhu Kaichuan, Zhang Jianhua, Liu Shide. Advances in yeast transcription factor Gal4 [J]. Chinese Journal of Bioengineering, 2011 1 1 (01): 81-85.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 2452
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 669
  Illegal XhoI site found at 2430
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 286
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 2281