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Team:Groningen/Template/MODULE/Notebook/Bandage
From 2014.igem.org
Notebook
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The Bandage
June 30 - July 4
Freeze drying L. lactis strain NZ9800 and NZ9700
These cells were stored at -80 °C
July 7 - July 11
Growing an over night culture of the freeze dried
The cells grew on the plate, meaning that L. lactis is able to survive the freeze drying
July 21- July 25
Growing a new overnight cultures and preparing the chemicals for the first polyacrylamide gels with L. lactis cells
Pouring a polyacrylamide hydrogel with Lactococcus lactis NZ9700 in it
Four polyacrylamide gels with different concentration were poured, one of 10 %, 15 %, 20 %, 25 %, and 30 % acrylamide, we decided to continue with 20 % for now
Pouring a polyacrylamide gel containing an overnight culture, this gel is divided in four, two gels were freeze dried, and two more were incubated overnight at 30 °C with fresh M17 medium
These gels were checked by phase contrast microscopy after incubation overnight, a lot of cells were visible, but we could not distinguish the living of dead bacteria, therefore we decided to continue the experiment with an inducable GFP expressing strain
July 28 - August 1
Preparing a GFP stock of L. lactis NZ9000 with pNZ8048g, and growing an overnight culture of this strain
Inducing the L. lactis NZ9000 with pNZ8048g with different concentrations of ZnSO4, the cells were observed under the microscope and some GFP expression was observed
Figure 1:
The L. lactis NZ9000 strain with pNZ8048g under the microscope, on the left the GFP channel is shown, and on the right the ordinary phase contrast picture is displayed
4 August - 8 August
This time we'll try pouring a gel on ice. This is because during the solidifying of acrylamid there is a lot of heat, we were a bit scared this heat could destroy our cells.
Unfortunately the solidifying process will slow down to a point that it will take more then an hour to complete. We've decided that this wouldn't work, so we continue without using ice.
Poured a gell with the GFP strain. Freeze a half, and the other half was put into the stove.
The next day these were both incubated with ZnSO4 for an hour. After an hour of incubating this didn't give any results. Incubation time should be longer the next time.
Figure 2:
The L. lactis NZ9000 strain with pNZ8048g under the microscope, on the left the GFP channel is shown, and on the right the phase contrast picture is displayed
August 11 - August 15
A polyacrylamide gel with the L. lactis NZ9000 strain with pNZ8048g, we incubated the gels in GM17 media with ZnSO4 for two hours at 30 °C, and observed the gels under the microscope
Some GFP expressing L. lactis was seen under the microscope
Repeating the experiment looking for the best conditions
Figure 3:
The L. lactis NZ9000 with pNZ8048g, seen through a phase contrast microscope, on the right the GFP channel is displayed
August 18 - August 22
Pouring a polyacrylamide gel with an overnight culture of L. lactis NZ9000 with pNZ8048g, the gel was split into several parts, and the parts were observed after certain time-spams divided over two weeks
L. lactis NZ9000 with pNZ8048g seems to be able to survive the polyacrylamide is liquid state and the polymerization process
25 August - 29 August
test 1
test 2
test 3
