Team:TMU-Tokyo/Notebook/Assay 1 Protocol and Result

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<Br>
<Br>
<B>■Assay</B><Br>
<B>■Assay</B><Br>
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Device1 Assay<Br>
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<A Href="https://2012.igem.org/Team:TMU-Tokyo/Notebook/Assay_0_Protocol_and_Result">Assay0</A><Br>
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Device2 Assay<Br>
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Device1 Assay</A><Br>
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Device3 Assay<Br>
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<A Href="https://2012.igem.org/Team:TMU-Tokyo/Notebook/Assay_2_Protocol_and_Result">Device2 Assay<Br>
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<A Href="https://2012.igem.org/Team:TMU-Tokyo/Notebook/Assay_3_Protocol_and_Result">Device3 Assay</A><Br>
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<p class="description">
<p class="description">
<b>■Assay1 Protocol</b><Br>
<b>■Assay1 Protocol</b><Br>
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<Br><Br>
 
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I. Assay of resistance for formaldehyde<Br>
 
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1. Make mediums<Br>
 
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Agar medium's recipe<Br>
 
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Agar mediums<Br>
 
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Dissolve parl  core agar medium 37.5 g into DW 200 ml.<Br>
 
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Autoclave for 20 minutes at 121 °C.<Br>
 
<Br>
<Br>
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Solution mediums<Br>
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1. In LB medium (liquid), wound bacteria frm-GFP, were incubated for 6 hours at room shaking culture.<Br>
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Dissolve glucose 2 g, polypeptone 2 g and yeast extract 2 into DW 200 ml.<Br>
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2. Was placed 5μl (0.04%) formalin solution to take the sample tube 500μl culture was diluted 200-fold there.<Br>
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Autoclave for 20 minutes at 121 °C.<Br>
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3.4 Degrees, I was centrifuged for 2 minutes at 120rpm<Br>
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4. Supernatant was removed, I was vortexed<Br>
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5. Put on the preparations taking 3μl from the tube, I observed a cover glass.<Br>
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6. I observed as a control those that do not contain formaldehyde.<Br>
<Br>
<Br>
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Phosphate buffer<Br>
 
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Make disodium hydrogen-phosphate aq 300 ml (0.2 M) and sodium<Br>
 
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dihydrogenphosphate dihydrate aq 300 ml (0.2 M) for pH 6.8.<Br>
 
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<Br>
 
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2. Cultivate E. coli in solution mediums for overnight.<Br>
 
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3. Prepare 1ml 0.1 %, 1 %, 5 % and 10 % solution of formaldehyde and formate.<Br>
 
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4. Add 1 μl E. coli into these solutions for 10 minutes.<Br>
 
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5. Plate on an agar medium.<Br>
 
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6. Incubate at 37 °C for overnight.<Br>
 
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<Br>
 
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II. Additional experiment for formaldehyde<Br>
 
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1. Prepare 1 ml 0.04 %, 0.08 %, 0.12 %, 0.16 % and 0.2 % of
 
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formaldehyde solution.<Br>
 
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2. Add 1 μl E. coli into these solutions for 10 minutes.<Br>
 
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3. Plate on an agar medium.<Br>
 
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4. Incubate at 37 °C for overnight.<Br>
 
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<p class="description">
<p class="description">
<b>■Assay1 Result</b><Br>
<b>■Assay1 Result</b><Br>
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GFP expression was not observed either.
 +
50 times, 100 times, 400 times, 600 times, 800 times, created in 1000-fold diluted solution of the same procedure, but I was the same experiment, the expression of GFP was not observed either.
</p>
</p>

Latest revision as of 02:26, 27 September 2012

 




■Experiment
1st week (8.13 - 8.19)
2nd week (8.20 - 8.26)
3rd week (8.27 - 9. 2)
4th week (9. 3 - 9. 9)
5th week (9.10 - 9.16)
6th week (9.17 - 9.23)
7th week (9.24 - 9.30)


■Protocols


■Assay
Assay0
Device1 Assay
Device2 Assay
Device3 Assay




Assay 1


■Assay1 Protocol

1. In LB medium (liquid), wound bacteria frm-GFP, were incubated for 6 hours at room shaking culture.
2. Was placed 5μl (0.04%) formalin solution to take the sample tube 500μl culture was diluted 200-fold there.
3.4 Degrees, I was centrifuged for 2 minutes at 120rpm
4. Supernatant was removed, I was vortexed
5. Put on the preparations taking 3μl from the tube, I observed a cover glass.
6. I observed as a control those that do not contain formaldehyde.

■Assay1 Result
GFP expression was not observed either. 50 times, 100 times, 400 times, 600 times, 800 times, created in 1000-fold diluted solution of the same procedure, but I was the same experiment, the expression of GFP was not observed either.



 

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