Checking Genetic Fence Plasmids
- Running digests to confirm the status of biobrick plasmids from igem team fence over the summer and into the fall. If the proper-sized cutout appears, I will keep the plasmid checked and discard the others so as to consolidate the materials in the fence boxes down to one or two boxes.
- digested each plasmid with EcoR1 and Pst1 to confirm the presence of the widest parts of the biobrick sites as well.
- used between 1 and 3μL of plasmid, depending on concentration, in an attempt to stay at reasonably concentrations of DNA.
First (top) Row
| Number:
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
9
|
10
|
11
|
12
|
13
|
14
|
15
|
16
|
17
|
18
|
| Name:
|
KB+ Ladder
|
Barstar 1
|
Barstar 2
|
RXRHm 2
|
RXRHm 1
|
LacIN 2
|
Gal4 DBD 1
|
Gal4 DBD 2
|
VP16GalEcR 1
|
VP16GalEcR 2
|
GalEcR 1
|
GalEcR 2
|
LacIN 1
|
5xGalUAS 1
|
5xGalUAS 2
|
LacIN Nost Act2LacO 1
|
LacIN Nost Act2LacO 2
|
KB+ Ladder
|
| Expected Length(bp):
|
|
273
|
273
|
600+
|
600+
|
1119
|
438
|
438
|
714
|
714
|
1446
|
1446
|
1119
|
93
|
93
|
2657
|
2657
|
|
| Correct Size:
|
|
yes +
|
yes +
|
yes +
|
yes +
|
yes +
|
yes
|
yes +
|
yes
|
yes
|
short
|
yes
|
blank
|
not visible +
|
not visible +
|
no band visible
|
blank
|
|
Second Row
| Number:
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
9
|
10
|
11
|
12
|
13
|
14
|
15
|
16
|
17
|
18
|
| Name:
|
KB+ Ladder
|
EcR
|
pENT
|
RXRLc
|
Act2LacO 1
|
VP16 1
|
35s 1
|
35s 2
|
Act2LacO 2
|
Barnase 1
|
Barnase 2
|
CaMV
|
Barstar Nost pENT 2
|
Barstar Nost pENT 1
|
|
|
|
KB+ Ladder
|
| Expected Length(bp):
|
|
1008
|
500
|
630
|
1282
|
276
|
56
|
56
|
1282
|
~250
|
~250
|
515
|
1032
|
1032
|
|
|
|
|
| Expected Length(bp):
|
|
yes +
|
yes +
|
yes +
|
yes, but with a second shorter band
|
yes +
|
yes +
|
yes +
|
yes, but with a second shorter band
|
yes +
|
yes +
|
yes +
|
yes +
|
yes +
|
|
|
|
|
- each successive image follows the gel running an additional 15 minutes after the initial 15 minute run
- Summary: at least one copy of each part is good except for Act2LacO (which has the same old problem with the weird two bands) and LacIN-Nost-Act2LacO, which probably is a result of the fact that the LacO from which it was cloned has that weird band (and did at the time of cloning as well)
To Do
- do a series of diagnostic digests on Act2LacO with different enzymes and also without any restriction enzymes
- based on the results of that, prepare to re-ligate LacIN-Nost-Act2LacO (which I need as a whole part to do gibson on the whole fence)
|
Project name
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