Notebook > AND Gate 1 > Output > Min Lin's Notes
2009.9.15
Transformation from the part distribution the P2 activator PhiR73 delta and one of its promoter PO.
Plate overnight.
2009.9.16
Pick one of the PhiR73 delta colony and PO promoter colony, shake in the incubator.
Miniprep PhiR73 delta and PO promoter plasmid.
2009.9.17
Enzyme Digestion of the PO promoter plasmid:
| SpeI | 1ul
|
| PstI | 1ul
|
| Buffer 3 | 2ul
|
| Plasmid | 3ul
|
| ddH2O | 13ul
|
Enzyme Digestion of the E0840
| XbaI | 1ul
|
| PstI | 1ul
|
| Buffer | 2ul
|
| Plasmid | 5ul
|
| ddH2O | 11ul
|
GEL assessment of the PO promoter plasmid(SP digest)
CIAP it for 20 min.
Directly purify it.
GEL: Enzyme Digestion of E0840, gel purification of the insert.
Ligation:
PO promoter vector 1ul
E0840 insert 7ul
Ligase 1ul
Ligation buffer 1ul
Transformation.
2009.9.18
Pick single colony of the PO-GFP and PCR to assess whether it is correct.
All 5 of the colonies are correct.
Shake in the incubator.
Miniprep.
Disgest PO-GFP with XbaI and PstI, overnight.
Get the Salicylate inducible promoter from zgs and digest with speI and pstI, overnight.
2009.9.19
GEL:
PO-GFP and get the XP insert.
Sal promoter and pointmutation colonies for assessment.
Send pointmutation plasmid for sequencing.
CIAP the sal promoter vector for 20 min
Ligation:
PO-GFP XP insert and sal promoter vector
Transformation.
2009.9.20
Pick colonies of the sal-PO-GFP
Colony PCR to assess. (get the correct colony, shake in the incubator)
Miniprep.
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