Team:Paris/Modeling/More The Steps

• 1st step: Introduce the tested promoter:

-Cut Promoter with EcoRI and SpeI -Cut Principal plasmid with EcoRI and XbaI

• 2nd step: Introduce the 1st tested transcription factor:

-Cut Gene with XbaI and SpeI -Cut Principal plasmid+Promoter with SpeI -Check the right orientation of the 1st gene by PCR with appropiated primers

If wanted…

• 3rd step: Introduce the 2nd gene into the Accessory plasmid

-Cut Gene with XbaI and SpeI -Cut Accessory Plasmid with XbaI and SpeI -Check the right orientation of the 2nd gene by PCR with appropiated primers4

• 4th step: Introduce the 2nd gene expression system into the Principal Plasmid:

-Cut Accessory Plasmid+2nd gene with PstI -Cut Principal plasmid+Promoter+1st gene with SpeI -Check the right orientation of the 2nd gene expression system by PCR with appropiated primers