Part:BBa_K2100044

pEXPR pERE6:BM3R1

This construct is an expression vector of a pERE6 promoter upstream of a BM3R1 repressor. pEREx6 is a synthetic mammalian promoter that responds to estrogen receptor activated by estradiol (E2). pEREx6 consists of six 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 18
  Illegal EcoRI site found at 333
  Illegal EcoRI site found at 343
  Illegal EcoRI site found at 749
  Illegal XbaI site found at 42
  Illegal PstI site found at 566
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 18
  Illegal EcoRI site found at 333
  Illegal EcoRI site found at 343
  Illegal EcoRI site found at 749
  Illegal NheI site found at 983
  Illegal PstI site found at 566
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 18
  Illegal EcoRI site found at 333
  Illegal EcoRI site found at 343
  Illegal EcoRI site found at 749
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 18
  Illegal EcoRI site found at 333
  Illegal EcoRI site found at 343
  Illegal EcoRI site found at 749
  Illegal XbaI site found at 42
  Illegal PstI site found at 566
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 6
  Illegal EcoRI site found at 18
  Illegal EcoRI site found at 333
  Illegal EcoRI site found at 343
  Illegal EcoRI site found at 749
  Illegal XbaI site found at 42
  Illegal PstI site found at 566
  Illegal NgoMIV site found at 488
  Illegal NgoMIV site found at 723
  Illegal AgeI site found at 501
  
• 1000
  COMPATIBLE WITH RFC[1000]