Template:Team:Groningen/CONTENT/LOGBOOK/Grow a Biofilm on a Carrier Material
Grow a Biofilm on a Carrier Material
Bacteria are triggered by their environment to change their behavior. Bacteria, like B. subtilis, preferably grow on rich media. However, when the medium where they grow on is less rich in nutrients, they start to form biofilms. In our project we need a carrier material to grow our biofilm on to make it more stable. In this experiment B. subtilis was grown on different media and different carrier materials.
Find the best carrier material and medium for the biofilm of B. subtilis 3610 ComI.
Biofilm growth thrives best in a minimal medium like MSgg. LB medium is too rich with nutrients for the bacteria to trigger solid biofilm growth. The biofilms seen on LB are not more than a smear of bacteria. Biofilms grown on MSgg show more structural integrity than biofilms grown on LB. Although the biofilm on the Nitrocellulose 0.025µM looks nice, the biofilms on the carrier materials with larger pores (0.2µM-0.45µM) are more suited for our project since these biofilms are interwoven with the carrier.
<a class="postscriptum protocol" href="https://2015.igem.org/Team:Groningen/Protocols_and_Protocols/No_protocol">Finibus sapien, nec cursus...</a>
00:00, 25 March 2015 - 00:00, 26 March 2015
Plate 10 µL B. subtilis 3610 ComI overnight culture (OD: 2,28 (600nm) after 22h growth) on different carrier materials on LB agar and MSgg agar plates. MSgg was prepared according to protocol.
Carrier materials using LB agar as a medium
Nylon positive charge (Boeringer, 1269299)
Nytran Plus 0.45 µm (Schleicher en Schuell, AH0495-1)
PVDF (Rocke, 03010040001)
Nitrocellulose 0.2 µm (invitrogen, LC2000)
Nylon 0.45 µm (Merck Milipore, HNWP04700)
Nitrocellulose 0.025 µm (Merck Milipore, VSWP02500)
Control (no carrier)
Carrier materials using MSgg agar as a medium
Nytran Plus 0.45 µm (Schleicher en Schuell, AH0495-1)
PVDF (Rocke, 03010040001)
Nitrocellulose 0.2µm (invitrogen, LC2000)
Nylon 0.45 µm (Merck Milipore, HNWP04700)
Nitrocellulose 0.025µm ( Merck Milipore, VSWP02500)
Control (no carrier)
All plates with carrier and bacteria were put in the incubator at 30°C.
00:00, 26 March 2015 - 00:00, 26 March 2015
After 24 hours, the biofilm growth was checked.
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LB plates after 24 hours. A is the control, B: Nitrocellulose 0.025µM, C: Nitrocellulose 0.2µM, D: Nylon 0.45µM, E: Nytran Plus 0.45µM, F: Nylon positive charge and G: PVDF. On all but the PVDF membrane are colonies growing.
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MSgg plates after 24 hours. A: control, B: Nitrocellulose 0.2µM, C: Nitrocellulose 0.025µM, D: Nytran Plus 0.45µM, E: PVDF and F: Nylon 0.45µM. Bacterial growth is seen on all samples, except for PVDF.
00:00, 30 March 2015 - 00:00, 30 March 2015
After 120 hours, the biofilm growth was checked. Biofilm growth thrives better on MSgg as can be seen on the images below. After 120 hours there is a clear difference visible between the LB and MSgg conditions. Furthermore, Nitrocellulose 0.2µM shows an nice and even distribution of biofilm and PVDF doesn't show biofilm growth at all on the MSgg plate.
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LB plates after 120 hours. A is the control, B: Nitrocellulose 0.025µM, C: Nitrocellulose 0.2µM, D: Nylon 0.45µM, E: Nytran Plus 0.45µM, F: Nylon positive charge and G: PVDF. Bacterial biofilm growth is seen on all the carrier materials.
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MSgg plates after 24 hours. A: control, B: Nitrocellulose 0.2µM, C: Nitrocellulose 0.025µM, D: Nytran Plus 0.45µM, E: PVDF and F: Nylon 0.45µM. Bacterial growth is seen on all samples, except for PVDF.