Test pMnt/Lac

pMnt/Lac is constitutively on but is repressed by the LacI protein. The promoter is also repressed by the Mnt protein. The addition of IPTG should repress the LacI repressor and glow in the absence of the Mnt protein.

To Test repression of promoter

Some promoter→RBS+LacI+t+t+pLac/Mnt→RBS+GFP ''does not glow''

I13453 → B0034 +LacI+TT+ K091104→E0240

Some promoter→RBS+Mnt+t+t+pLac/Mnt→GFP ''does not glow''

I13453 → B0034 +Mnt+TT+ K091104→E0240

Some promoter→RBS+LacI+RBS+Mnt+t+t+pLac/Mnt→GFP ''does not glow''

I13453 → B0034 +LacI+RBS+Mnt+TT+ K091104→E0240

To Test the promoter and addition of IPTG

add IPTG

Some promoter→IPTG+RBS+LacI+t+t+pLac/Mnt→GFP ''glows''

I13453 → IPTG+B0034+LacI+TT+K091104→E0240

Some promoter→IPTG+RBS+LacI+Mnt+t+t+pLac/Mnt→GFP '' glows''

I13453 →IPTG+ B0034 +LacI+RBS+Mnt+TT+ K091104→E0240

LacI	Mnt	IPTG + GFP	IPTG - GFP
0	0	1	1
0	1	0	0
1	0	1	0
1	1	0	0

Steps to Build Hybrid Promoter

6/30/08

Perform a Ligation of Mnt/Lac (K091104) with E0240 (RBS + GFP)

Transform ligation over night

7/1/08

Clones were picked and set over night in LB + AMP