From 2009.igem.org
Wet
GVP Cluster
Discussion:
All colonies (transformants vector + promoter ligated with gvp) and o/n culture became red, so probably RFP is still in there...
What might be the problem? The vector with promoter self-ligated due to uncomplete digestion (done 15 July 2009) leading to the following fragments:
- vector+RFP linear
- vector
- RFP
When the upper fragment (~2kb) is contaminated with linear vector this would lead high efficiency self-ligation instead of ligating gvp with the emtpy vector.
- →TODO Therefore the restriction/purification/ligation will be redone and selection of non-red colonies will be made!
- →DONE Also plasmid from the o/n cultures will be purified and analysed by restriction analysis.
- Used Sigma Plasmid isolation kit.
- DNA concentrations:
Transporters
PCR 16-07
Component | B1 | B2 | C1 | C2 | A1 | A2 | E1 | E2 | F1 | F2
|
MM | 21 | 22 | 21 | 22 | 21 | 22 | 21 | 22 | 21 | 22
|
F 1uL | Fw | Fw | Fw | Fw | mut1 | mut1 | Rev | Rev | Fw | Fw
|
R 1uL | Rev | Rev | mut2 | mut2 | PCR2 | PCR2 | mut1 | mut1 | mut1RC | mut1RC
|
DNA 1uL | 1 | 0 | 1 | 0 | 1 | 0 | 1 | 0 | 1 | 0
|
taq 1uL | 1 | 1 | 1 | 1 | 1 | 1 | 1 | 1 | 1 | 1
|
|
|}
Metal Accumulation
Vectors
Dry