Difference between revisions of "Team:Nagahama/awarming"
(→Swarming Support) |
(→We helped Tokyo_Tech.) |
||
| Line 5: | Line 5: | ||
==We helped Tokyo_Tech.== | ==We helped Tokyo_Tech.== | ||
| − | + | ===Abstract=== | |
We helped team Tokyo_Tech by advising these 2 about chemotaxis experiments. | We helped team Tokyo_Tech by advising these 2 about chemotaxis experiments. | ||
| Line 12: | Line 12: | ||
Also, we gave Tokyo_Tech the raw data along with the protocol of the experiments for assaying the chemotactic activity of a wild type E.coli. | Also, we gave Tokyo_Tech the raw data along with the protocol of the experiments for assaying the chemotactic activity of a wild type E.coli. | ||
| − | + | ===Method=== | |
| − | 5 × M9 salts stock solution | + | ====5 × M9 salts stock solution==== |
6.4 ％ (w/v) Na2HPO4 × 7H2O | 6.4 ％ (w/v) Na2HPO4 × 7H2O | ||
1.5 ％ (w/v) KH2PO4 | 1.5 ％ (w/v) KH2PO4 | ||
| Line 20: | Line 20: | ||
0.5 ％ (w/v) NH4Cl | 0.5 ％ (w/v) NH4Cl | ||
| − | M9 swarming agar | + | ====M9 swarming agar==== |
1.25 ％ (v/v) glycerol | 1.25 ％ (v/v) glycerol | ||
20 ％ (v/v) 5 × M9 salts stock solution | 20 ％ (v/v) 5 × M9 salts stock solution | ||
| Line 28: | Line 28: | ||
0.3 ％ (w/v) agar | 0.3 ％ (w/v) agar | ||
| − | tripton broth | + | ====tripton broth==== |
1.0 ％ (w/v) tripton | 1.0 ％ (w/v) tripton | ||
1.0 ％ (w/v) NaCl | 1.0 ％ (w/v) NaCl | ||
| − | Swarming Assay | + | ====Swarming Assay==== |
We assayed chemotaxis of E. coli against cadmium and aspartic acid on soft agar, containing M9 synthetic medium, on that plate E. coli can swim. | We assayed chemotaxis of E. coli against cadmium and aspartic acid on soft agar, containing M9 synthetic medium, on that plate E. coli can swim. | ||
Protocol | Protocol | ||
| Line 41: | Line 41: | ||
5.The plate was incubated at 30℃. | 5.The plate was incubated at 30℃. | ||
| − | + | ===Result and Discussion=== | |
[[File: Asp_swamming.png|300px]] | [[File: Asp_swamming.png|300px]] | ||
Revision as of 03:19, 16 September 2015
Contents
Swarming Support
We helped Tokyo_Tech.
Abstract
We helped team Tokyo_Tech by advising these 2 about chemotaxis experiments. 1. In order to increase the chemotactic activity of E. coli, the best concentration of agar is 0.3 %. 2. It is better to directly stick the chip inside the agar and then inject the E .coli, than to place the E. coli on top of the agar Also, we gave Tokyo_Tech the raw data along with the protocol of the experiments for assaying the chemotactic activity of a wild type E.coli.
Method
5 × M9 salts stock solution
6.4 ％ (w/v) Na2HPO4 × 7H2O 1.5 ％ (w/v) KH2PO4 0.25 ％ (w/v) NaCl 0.5 ％ (w/v) NH4Cl
M9 swarming agar
1.25 ％ (v/v) glycerol 20 ％ (v/v) 5 × M9 salts stock solution 0.1 ％ (v/v) of CaCl2 × 2H2O stock solution (20 mg/mL) 0.1 ％ (v/v) MgSO4 stock solution (0.12 g/mL) 0.03 ％ (w/v) thiamine 0.3 ％ (w/v) agar
tripton broth
1.0 ％ (w/v) tripton 1.0 ％ (w/v) NaCl
Swarming Assay
We assayed chemotaxis of E. coli against cadmium and aspartic acid on soft agar, containing M9 synthetic medium, on that plate E. coli can swim. Protocol 1.E. coli JM109 was cultured at 30℃ for 12 hours with shaking (50 rpm). 2.Aliquot of the culture was spotted on the center of agar plate. 3.10 mM L-aspartic acid (40 μl) or 100 mM cadmium chloride (4 μl) was spotted on 25 mm distant from the center of the agar plate. 4.The plate was standed for 5 min at RT. 5.The plate was incubated at 30℃.
Result and Discussion
fig1：E. coli swarming to aspartic acid L: aspartic acid (10 mM, 40 μl) R: ddH2O (40 μl) Incubation: 108 hours temperature: 30 ℃ E. coli cultured was spotted on the center of agar plate, and chemotaxis was spotted on 25 mm distant from the center. The swarming circle of aspartic acid (L) was larger than that of ddH2O (R), and the swarming area around aspartate acid (L) was more larger than that of ddH2O (R), suggesting that E. coli swarming to aspartate acid. This experiment indicate that E. coli might have positive chemotaxis for aspartic acid.
fig2：E. coli hate cadmium. L: cadmium(100 mM, 4 μl) R: ddH2O (40 μl) Incubation: 110 hours temperature: 30 ℃ E. coli cultured was spotted on the center of agar plate, and chemotaxis was spotted on 25 mm distant from the center. The swarming circle of cadmium (L) was smaller than that of ddH2O (R), and the swarming area around cadmium (L) was more smaller than that of ddH2O (R), suggesting that E. coli hate cadmium. This experiment indicate that E. coli might have negative chemotaxis for cadmium.