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Team:Paris Bettencourt/Notebook/Foot Odor
From 2014.igem.org
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<p>No colonies in LBA + EM. Colonies observed in LBA w/o EM | <p>No colonies in LBA + EM. Colonies observed in LBA w/o EM | ||
</p> | </p> | ||
| + | <h5>27-06-2014</h5> | ||
| + | <h6>To prepare LBA and LB broth for B.Subtilis mutants</h6> | ||
| + | <h6>Procedure</h6> | ||
| + | <p>We added 2.5 ul of the EM (20mg/ml) to 50ml of LB broth. We added 25 ul of EM to 250 ml of LBA and prepared the plates & stored them at 4° C.</p> | ||
| + | <h5>30-06-2014</h5> | ||
| + | <h6>Goal: To prepare 10% glycerol stock of wt and mutant (BKE24040, BKE24050, BKE24080) B.Subtilis </h6> | ||
| + | <h6>Procedure</h6> | ||
| + | <p>We inoculated Single colony of both wt (1A1) and mutant (BKE24040, BKE24050, BKE24080) in 5ml of LB broth supplemented w/o EM for wt and with EM for mutants. Incubated the colonies @ 37°C overnight. After the overnight incubation the tubes were stirred well. 750 ul of the culture is transferred to cryogenic tube and 750 ul of 60% glycerol is added to the culture to obtain 10% glycerol stock. | ||
| + | </p> | ||
| + | <TABLE BORDER> | ||
| + | <tr> | ||
| + | <td colspan="1" rowspan="1"><p>Entry</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>Strain</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>Resistance</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>Source</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>Strain</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>Glycerol Stock Label</p></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1" rowspan="1"><p>sPB.011</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BKE24040</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>EM</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BGSC</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>B.subtilis with 2-oxoisovalerate dehydrogenase beta subunit knockout</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>G.11</p></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1" rowspan="1"><p>sPB.012</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BKE24050</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>EM</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BGSC</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>B.subtilis with 2-oxoisovalerate dehydrogenase alpha subunit knockout</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>G.12</p></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1" rowspan="1"><p>sPB.013</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BKE24080</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>EM</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BGSC</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>B.subtilis with leucine dehydrogenase knockout</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>G.13</p></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1" rowspan="1"><p>sPB.014</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>1A1</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>-</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>BGSC</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>B.subtilis wt strain</p></td> | ||
| + | <td colspan="1" rowspan="1"><p>G.14</p></td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | <p> The cryo tubes were stored at -20°C.</p> | ||
</br> | </br> | ||
</div> | </div> | ||
Revision as of 13:01, 20 August 2014
Paris Bettencourt 2014
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Foot Odor
June
23-06-2014
Goal: To obtain single colonies of both Wt and mutant B.Subtilis
We received the wt (1A1) and mutant BKE24040 (isovalerate dehydrogenase beta KO), BKE24050 (isovalerate dehydrogenase alpha KO), BKE24080 (Leucine dehydrogenase KO) from BGSC on 23-06-2014. The wt strain (1A1) was on a paper strip. We received 3 strips from BGSC (Bacillus genetic stock centre). Where as the mutant strains arrived on perti-plates.
Procedure
We plated two strips in two different LBA plates. The plates were free from antibiotics. We placed the strips gently on the LBA and added 1 drop of sterile H2O. After few seconds we streaked the paper strip on the LBA and discarded the strip. The LBA plates were incubated at 37° C. We took single colony from the mutant B.subtilis petri-plates and streaked them on LBA+EM petri plates.
Results
Good growth of both the wt and mutant B.subtilis were observed followed by the overnight incubation
24-06-2014
To obtain single colonies of Wt b.Subtilis from 23-06-14 plate
Procedure
we took a single colony and streaked it on a new LBA with and w/o Erythromycin.
Results
No colonies in LBA + EM. Colonies observed in LBA w/o EM
27-06-2014
To prepare LBA and LB broth for B.Subtilis mutants
Procedure
We added 2.5 ul of the EM (20mg/ml) to 50ml of LB broth. We added 25 ul of EM to 250 ml of LBA and prepared the plates & stored them at 4° C.
30-06-2014
Goal: To prepare 10% glycerol stock of wt and mutant (BKE24040, BKE24050, BKE24080) B.Subtilis
Procedure
We inoculated Single colony of both wt (1A1) and mutant (BKE24040, BKE24050, BKE24080) in 5ml of LB broth supplemented w/o EM for wt and with EM for mutants. Incubated the colonies @ 37°C overnight. After the overnight incubation the tubes were stirred well. 750 ul of the culture is transferred to cryogenic tube and 750 ul of 60% glycerol is added to the culture to obtain 10% glycerol stock.
Entry |
Strain |
Resistance |
Source |
Strain |
Glycerol Stock Label |
sPB.011 |
BKE24040 |
EM |
BGSC |
B.subtilis with 2-oxoisovalerate dehydrogenase beta subunit knockout |
G.11 |
sPB.012 |
BKE24050 |
EM |
BGSC |
B.subtilis with 2-oxoisovalerate dehydrogenase alpha subunit knockout |
G.12 |
sPB.013 |
BKE24080 |
EM |
BGSC |
B.subtilis with leucine dehydrogenase knockout |
G.13 |
sPB.014 |
1A1 |
- |
BGSC |
B.subtilis wt strain |
G.14 |
The cryo tubes were stored at -20°C.
July
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August
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September
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October
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