Difference between revisions of "Test pLsrA/cI"
| Line 6: | Line 6: | ||
'''1)Control group''' | '''1)Control group''' | ||
| − | plsrA/ | + | plsrA/cI→GFP glows |
| − | K091106+E0240 | + | [http://partsregistry.org/Part:BBa_K091106 K091106]+[http://partsregistry.org/Part:BBa_E0240 E0240] |
'''2) Test LsrR repression:''' | '''2) Test LsrR repression:''' | ||
| − | pBAD→RBS+LsrR+t+t+plsrA/ | + | pBAD→RBS+LsrR+t+t+plsrA/cI→GFP does not glow |
| − | I13453+B0034+K091001+B0015+K091106+E0240 | + | [http://partsregistry.org/Part:BBa_I13453 I13453]+[http://partsregistry.org/Part:BBa_B0034 B0034]+[http://partsregistry.org/Part:BBa_K091001 K091001]+[http://partsregistry.org/Part:BBa_B0015 B0015]+[http://partsregistry.org/Part:BBa_K091106 K091106]+[http://partsregistry.org/Part:BBa_E0240 E0240] |
add AI-2 | add AI-2 | ||
| − | pBAD→RBS+LsrR+t+t+plsrA/ | + | pBAD→RBS+LsrR+t+t+plsrA/cI→GFP glows |
| − | |||
| − | |||
| + | [http://partsregistry.org/Part:BBa_I13453 I13453]+[http://partsregistry.org/Part:BBa_B0034 B0034]+[http://partsregistry.org/Part:BBa_K091001 K091001]+[http://partsregistry.org/Part:BBa_B0015 B0015]+[http://partsregistry.org/Part:BBa_K091106 K091106]+[http://partsregistry.org/Part:BBa_E0240 E0240] | ||
'''3) Test cI repression:''' | '''3) Test cI repression:''' | ||
| Line 28: | Line 27: | ||
pBAD→RBS+cI+TT+plsrA/cI→GFP does not glow | pBAD→RBS+cI+TT+plsrA/cI→GFP does not glow | ||
| − | I13453+B0034+C0051+B0015+K091106+E0240 | + | [http://partsregistry.org/Part:BBa_I13453 I13453]+[http://partsregistry.org/Part:BBa_B0034 B0034]+[http://partsregistry.org/Part:BBa_C0051 C0051]+[http://partsregistry.org/Part:BBa_B0015 B0015]+[http://partsregistry.org/Part:BBa_K091106 K091106]+[http://partsregistry.org/Part:BBa_E0240 E0240] |
| − | |||
'''4) Test LsrR and cI repression:''' | '''4) Test LsrR and cI repression:''' | ||
| Line 35: | Line 33: | ||
pBAD→RBS+lsrR+cI+TT+plsrA/cI→GFP does not glows | pBAD→RBS+lsrR+cI+TT+plsrA/cI→GFP does not glows | ||
| − | I13453+B0034+K091001+C0051+B0015+K091106+E0240 | + | [http://partsregistry.org/Part:BBa_I13453 I13453]+[http://partsregistry.org/Part:BBa_B0034 B0034]+[http://partsregistry.org/Part:BBa_K091001 K091001]+[http://partsregistry.org/Part:BBa_C0051 C0051]+[http://partsregistry.org/Part:BBa_B0015 B0015]+[http://partsregistry.org/Part:BBa_K091106 K091106]+[http://partsregistry.org/Part:BBa_E0240 E0240] |
add AI-2 | add AI-2 | ||
| Line 41: | Line 39: | ||
pBAD→RBS+lsrR+cI+TT+plsrA/cI→GFP does not glows | pBAD→RBS+lsrR+cI+TT+plsrA/cI→GFP does not glows | ||
| − | I13453+B0034+K091001+C0051+B0015+K091106+E0240 | + | [http://partsregistry.org/Part:BBa_I13453 I13453]+[http://partsregistry.org/Part:BBa_B0034 B0034]+[http://partsregistry.org/Part:BBa_K091001 K091001]+[http://partsregistry.org/Part:BBa_C0051 C0051]+[http://partsregistry.org/Part:BBa_B0015 B0015]+[http://partsregistry.org/Part:BBa_K091106 K091106]+[http://partsregistry.org/Part:BBa_E0240 E0240] |
{| border="1" cellpadding="2" | {| border="1" cellpadding="2" | ||
Revision as of 15:07, 1 July 2008
Test pLsrA/cI promoter
The pLsrA/cI promoter is constitutively off and is repressed by the lsrR protein. The addition of AI-2 should de-repress the lsrR operon causing it to glow. The addition of the cI protein should repress the promoter, causing it to not glow.
1)Control group
plsrA/cI→GFP glows
2) Test LsrR repression:
pBAD→RBS+LsrR+t+t+plsrA/cI→GFP does not glow
I13453+B0034+K091001+B0015+K091106+E0240
add AI-2
pBAD→RBS+LsrR+t+t+plsrA/cI→GFP glows
I13453+B0034+K091001+B0015+K091106+E0240
3) Test cI repression:
pBAD→RBS+cI+TT+plsrA/cI→GFP does not glow
I13453+B0034+C0051+B0015+K091106+E0240
4) Test LsrR and cI repression:
pBAD→RBS+lsrR+cI+TT+plsrA/cI→GFP does not glows
I13453+B0034+K091001+C0051+B0015+K091106+E0240
add AI-2
pBAD→RBS+lsrR+cI+TT+plsrA/cI→GFP does not glows
I13453+B0034+K091001+C0051+B0015+K091106+E0240
| LsrR | cI | AI-2 + GFP | AI-2 - GFP |
|---|---|---|---|
| 0 | 0 | 1 | 1 |
| 1 | 0 | 1 | 0 |
| 0 | 1 | 0 | 0 |
| 1 | 1 | 0 | 0 |
Ligation plan for three test sequences
1)
K091106 ligated with E0240
need to ligate some promoter (digest with E/S) with an RBS (digest E/X) and ligate lsrR(digest with E/S) with a double terminator (digest with E/X)
2)
K091106 ligated with E0240
need to ligate some promoter (digest with E/S) with an RBS (digest E/X) and ligate lsrR(digest with E/S) with a double terminator (digest with E/X)
then ligate the promoter+RBS to the 5' end of the lsrR+TT parts together
then ligate that part 5' to the plsrA/cI+GFP
3)
ligate cI to the double terminator
the ligate the promoter+RBS from the first test sequence to the 5' end of the cI+TT
ligate this to the 5' end of the plsrA/cI+GFP
4)
ligate the cI+TT from the second test sequence to the 3' end of the lsrR coding region
then ligate the promoter+RBS from the first test sequence to the 5' end of that
next ligate the plsrA/cI to the 3' end of that entire sequence
the 2) and 4) test sequences will also need to be tested by adding AI-2
the 2) test sequence should glow and the 4) should not.
