Part:BBa_K2100043

pEXPR pERE5:BM3R1

This construct is an expression vector of a pERE5 promoter upstream of a BM3R1 repressor. pEREx5 is a synthetic mammalian promoter that responds to estrogen receptor activated by estradiol (E2). pEREx5 consists of five repeats of the estrogen response element (ERE) with the sequence 5′-GGTCAnnnTGACC-3 upstream of a minimal promoter (minCMV). We interspaced these elements with 22 randomly selected bases in between, the same spacing between TREs in the TRE promoter. This design choice was made because when an ERE is not properly oriented, it has trouble interacting with ERα and polymerases. Since we wanted to decrease the probability of helical turns inhibiting the transcriptional activity of promoters, we modeled the spacing after the TRE promoter.

In endometrial cells, estrogen receptors facilitate the cell's response to estrogen, initiating the proliferative phase of the menstrual cycle. Mechanistically, estrogen diffuses through the cell membrane and binds to the ligand binding domain of the ER. The DNA-binding domain of the estrogen receptor becomes exposed. The receptor then relocalizes to the nucleus from the cytoplasm, recruits co-activators, and acts as a transcription factor for the BM3R1 gene. When transcribed, BM3R1 represses the DNA sequences found on the pBM3R1 promoter with the help of the corepressor VP16-Gal4.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 271
  Illegal EcoRI site found at 281
  Illegal EcoRI site found at 687
  Illegal PstI site found at 504
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 271
  Illegal EcoRI site found at 281
  Illegal EcoRI site found at 687
  Illegal NheI site found at 921
  Illegal PstI site found at 504
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 271
  Illegal EcoRI site found at 281
  Illegal EcoRI site found at 687
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 271
  Illegal EcoRI site found at 281
  Illegal EcoRI site found at 687
  Illegal PstI site found at 504
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 271
  Illegal EcoRI site found at 281
  Illegal EcoRI site found at 687
  Illegal PstI site found at 504
  Illegal NgoMIV site found at 426
  Illegal NgoMIV site found at 661
  Illegal AgeI site found at 439
  
• 1000
  COMPATIBLE WITH RFC[1000]