Team:UNICAMP-EMSE Brazil/Notebook/29 August 2011
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29 August 2011
Finishing the first two biobricks!!!
Objective:
- Checking digestion of yesterday's ligation with EcoRI and PstI
Samples:
- Gel 1:
- HlyA+Terminator C1/ HlyA+Terminator C2/ HlyA+Terminator C3/ HlyA+Terminator C4/ Const. Promoter+ RBS+SoxR+ Terminator C1/ Const. Promoter+ RBS+SoxR+ Terminator C2/ SoxS+RBS+IL 10 C1/ SoxS+RBS+IL 10 C2/ flHDC+IL 12 C1/ flHDC+IL 12 C2/ RBS+HlyD+TolC+Terminator C2a/ RBS+HlyD+TolC+Terminator C1a
- Gel 2:
- SoxS+RBS+GFP C1/ SoxS+RBS+GFP C2/ RBS+HlyD+TolC+Terminator C1/ RBS+HlyD+TolC+Terminator C2/ flHDC+RBS+HlyB C1/ flHDC+RBS+HlyB C2/ SoxS+RBS+HlyB C1/ SoxS+RBS+HlyB C2/ flHDC+RBS+GFP C1/ flHDC+RBS+GFP C2/ Constitutive Promoter+ RBS+QseB+RBS+QseC+Terminator C1/ Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C2/ Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C3/ Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C4
- Ladder:
- 100-10.000 bp (Fermentas)
- Gel Agarose concentration:
- 1,5%
- Results:
Gene | Total size (gene+vector) | Linear vector size | Gene size | Result |
---|---|---|---|---|
HlyA+Terminator C1 | 3527 bp | 3205 bp | 322 bp | No visible band |
HlyA+Terminator C2 | 3527 bp | 3205 bp | 322 bp | OK |
HlyA+Terminator C3 | 3527 bp | 3205 bp | 322 bp | OK |
HlyA+Terminator C4 | 3527 bp | 3205 bp | 322 bp | OK |
Const. Promoter+ RBS+SoxR+ Terminator C1 | 2732 bp | 2079 bp | 653 bp | Seems smaller |
Const. Promoter+ RBS+SoxR+ Terminator C2 | 2732 bp | 2079 bp | 653 bp | Seems smaller |
SoxS+RBS+IL 10 C1 | 3543 bp | 2800 bp | 743 bp | No visible bands |
SoxS+RBS+IL 10 C2 | 3543 bp | 2800 bp | 743 bp | OK |
flHDC+IL 12 C1 | 4584 bp | 2800 pb | 1784 | No visible bands |
flHDC+IL 12 C2 | 4584 bp | 2800 bp | 1784 | OK |
RBS+HlyD+TolC+Terminator C2a | 6376 bp | 3205 bp | 3171 bp | Run more |
RBS+HlyD+TolC+Terminator C1a | 6376 bp | 3205 bp | 3171 bp | Run more |
SoxS+RBS+GFP C1 | 3638 bp | 2800 bp | 838 bp | No visible band |
SoxS+RBS+GFP C2 | 3638 bp | 2800 bp | 838 bp | OK |
RBS+HlyD+TolC+Terminator C1 | 6376 bp | 3205 bp | 3171 bp | Run more |
RBS+HlyD+TolC+Terminator C2 | 6376 bp | 3205 bp | 3171 bp | Run more |
flHDC+RBS+HlyB C1 | 5109 bp | 2800 bp | 2309 bp | OK |
flHDC+RBS+HlyB C2 | 5109 bp | 2800 bp | 2309 bp | No visible band |
SoxS+RBS+HlyB C1 | 5086 bp | 2800 bp | 2286 bp | OK |
SoxS+RBS+HlyB C2 | 5086 bp | 2800 bp | 2286 bp | OK |
flHDC+RBS+GFP C1 | 3661 bp | 2800 bp | 861 bp | No visible bands |
flHDC+RBS+GFP C2 | 3661 bp | 2800 bp | 861 bp | No visible bands |
Constitutive Promoter+RBS+ QseB+RBS+QseC+Terminator C1 | 4295 bp | 2079 bp | 2216 bp | Run more |
Constitutive Promoter+RBS+ QseB+RBS+QseC+Terminator C2 | 4295 bp | 2079 bp | 2216 bp | Run more |
Constitutive Promoter+RBS+ QseB+RBS+QseC+Terminator C3 | 4295 bp | 2079 bp | 2216 bp | Run more |
Constitutive Promoter+RBS+ QseB+RBS+QseC+Terminator C4 | 4295 bp | 2079 bp | 2216 bp | Run more |
Same electrophoresys but after a longer period run:
Samples:
- Gel 3:
- HlyA+Terminator C1/ HlyA+Terminator C2/ HlyA+Terminator C3/ HlyA+Terminator C4/ Const. Promoter+ RBS+SoxR+ Terminator C1/ Const. Promoter+ RBS+SoxR+ Terminator C2/ SoxS+RBS+IL 10 C1/ SoxS+RBS+IL 10 C2/ flHDC+IL 12 C1/ flHDC+IL 12 C2/ RBS+HlyD+TolC+Terminator C2a/ RBS+HlyD+TolC+Terminator C1a
- Gel 4:
- SoxS+RBS+GFP C1/ SoxS+RBS+GFP C2/ RBS+HlyD+TolC+Terminator C1/ RBS+HlyD+TolC+Terminator C2/ flHDC+RBS+HlyB C1/ flHDC+RBS+HlyB C2/ SoxS+RBS+HlyB C1/ SoxS+RBS+HlyB C2/ flHDC+RBS+GFP C1/ flHDC+RBS+GFP C2/ Constitutive Promoter+ RBS+QseB+RBS+QseC+Terminator C1/ Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C2/ Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C3/ Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C4
- Ladder:
- 100-10.000 bp (Fermentas)
- Gel Agarose concentration:
- 1,5%
- Results:
Gene | Total size (gene+vector) | Linear vector size | Gene size | Result |
---|---|---|---|---|
RBS+HlyD+TolC+Terminator C2a | 6376 pb | 3205 pb | 3171 pb | Superposition of the bands, but wont be a problem in the next ligation, we will linearize only |
RBS+HlyD+TolC+Terminator C1a | 6376 pb | 3205 pb | 3171 pb | Superposition of the bands, but wont be a problem in the next ligation, we will linearize only |
RBS+HlyD+TolC+Terminator C1 | 6376 pb | 3205 pb | 3171 pb | Superposition of the bands, but wont be a problem in the next ligation, we will linearize only |
RBS+HlyD+TolC+Terminator C2 | 6376 pb | 3205 pb | 3171 pb | Superposition of the bands, but wont be a problem in the next ligation, we will linearize only |
Constitutive Promoter+ RBS+QseB+RBS+QseC+Terminator C1 | 4295 pb | 2079 pb | 2216 pb | Seems to be OK |
Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C2 | 4295 pb | 2079 pb | 2216 pb | Seems to be OK |
Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C3 | 4295 pb | 2079 pb | 2216 pb | Seems to be OK |
Const. Promoter+ RBS+QseB+RBS+QseC+Terminator C4 | 4295 pb | 2079 pb | 2216 pb | Seems to be OK |
OBS:-Inoculation of new colonies of flHDC+RBS+IL-12 and flHDC+RBS+GFP
Wowwww first two biobricks assembled !!! :) S2