Team:KAIST-Korea/Notebook/Diary/September

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== September 1 ==
== September 1 ==
pREP41 vector has arrived.
pREP41 vector has arrived.
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==September ? ==
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==September 2 ~ September 21 ==
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We designed PCR schedules and do PCR experiment. When PCR result was not satisfied, then we repeated PCR experiment. There are PCR details on <a href=https://2010.igem.org/Team:KAIST-Korea/Project/Methods/PCR_Result>PCR experiment menu</a>.  
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=== V_STAT1 (expression) ===
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<td>Primer</td><td>Direction</td><td>Sequences</td><td>Length</td>
 
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<td>V_STAT_F</td><td>Forward(NdeI)</td><td>5’-<font color=blue>TGTT</font><font color=red>CATATG</font><font color=blue>GCTA</font>ATGTCTCAGTGGTACGAACTTCAG-3’</td><td>24bp</td>
 
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<td>V_STAT_R</td><td>Reverse(BamHI)</td><td>5’-<font color=blue>TATC</font><font color=red>GGATCC</font><font color=blue>GAAT</font>TTACACTTCAGACACAGAAATCAAC-3’</td><td>25bp</td>
 
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<td><b>Materials</b></td><td><b>Volume</b></td>
 
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<td>STAT1(KRIBB) (template)</td><td></td>
 
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<td>V_STAT_F</td><td></td>
 
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<td>V_STAT_R</td><td></td>
 
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<td>dNTP (dNTP mixture 2.5mM TaKaRa)</td><td></td>
 
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<td>10x buffer (10x cloned Pfu Reaction buffer)</td><td></td>
 
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<td>DW (3rd sterile water)</td><td></td>
 
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<td>Total</td><td>50ul</td>
 
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[[Image:PCR_cycle.jpg]]
 
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=== STAT1 (submission) ===
 
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[[Image:PCR_STAT1.jpg]]
 
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<td>Primer</td><td>Direction</td><td>Sequences</td><td>Length</td>
 
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<td>STAT_F</td><td>Forward</td><td>5’-ATGTCTCAGTGGTACGAACTTCAG-3’</td><td>24bp</td>
 
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<td>STAT_R</td><td>Reverse</td><td>5’-TTACACTTCAGACACAGAAATCAAC-3’</td><td>25bp</td>
 
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<br>
 
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<table border=1>
 
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<td><b>Materials</b></td><td><b>Volume</b></td>
 
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</tr>
 
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<tr align=center>
 
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<td>STAT1(KRIBB) (template)</td><td></td>
 
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<tr align=center>
 
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<td>STAT_F</td><td></td>
 
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<td>STAT_R</td><td></td>
 
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<td>dNTP (dNTP mixture 2.5mM TaKaRa)</td><td></td>
 
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</tr>
 
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<tr align=center>
 
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<td>Taq (Pfu Turbo® DNA Polymerase 2.5U/ μl STRATAGENE)</td><td></td>
 
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</tr>
 
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<tr align=center>
 
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<td>10x buffer (10x cloned Pfu Reaction buffer)</td><td></td>
 
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</tr>
 
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<td>DW (3rd sterile water)</td><td></td>
 
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<td>Total</td><td>50ul</td>
 
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[[Image:PCR_cycle.jpg]]
 
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[[Image:PCR_FGFR.jpg]]
 
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[[Image:PCR_HR.jpg]]
 
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[[Image:PCR_GAS.jpg]]
 
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== September next ==
 
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Latest revision as of 07:21, 20 October 2010

 

September 1

pREP41 vector has arrived.

-> we will do E.coli transformation and increase its number.

pREP42-GFP vector has arrived (in E.coli)

-> will incubate a day and do spreading for further experiment.

Making culture media

  1. 500mL LB broth + agar 1.5% (7.5g)
  2. Do auto clave
  3. Cool it down until it reaches 50~60℃
  4. Add ampicillin(1000x 100mg/mL) and stir
  5. putting out bubbles on culture media
  6. pouling on petri dish and wait for 1 hour

Culture

  1. Competent cell + DN10A and leave it in ice for 30min
  2. Do heat shock for 45 sec and store it in ice for 2min
  3. Stabilize it in incubator for 30 to 40min
  4. Do spreading


September 2 ~ September 21

We designed PCR schedules and do PCR experiment. When PCR result was not satisfied, then we repeated PCR experiment. There are PCR details on PCR experiment menu.


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