Team:HokkaidoU Japan/Notebook/plastic Week 7

From 2012.igem.org

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<div class="hokkaidou-notebook-daily">
<div class="hokkaidou-notebook-daily">
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==August 17nd==
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===August 16th===
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<div>
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<div class="hokkaidou-section">
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===PCR===
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====PCR====
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<p>
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PCR of pGEM to get linear PhaA and PhaB using Laury primer and X-PhaA-up-F, S-PhaA-dw-R, X-PhaB-up-F, S-PhaB-dw-R.  
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A PCR of pGEM to get linear Pha A and PhaB using Laury primer.  
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{|class="hokkaidou-table-digestion"
{|class="hokkaidou-table-digestion"
   |-
   |-
-
   |pGEM(pGEM 1ul + DW 9ul)
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   |pGEM(pGEM 1 ul + DW 9 ul)
   |1 ul
   |1 ul
   |-
   |-
-
   |up primer(10mM)
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   |up primer(10 mM)
   |1 ul
   |1 ul
   |-
   |-
-
   |down primer(10mM)
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   |down primer(10 mM)
   |1 ul
   |1 ul
   |-
   |-
-
　|KOD plus Neo
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  |25 mM MgSO4
-
　|1ul
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  |3 ul
   |-
   |-
-
   |25mM MgSO4
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   |2 mM sNTPs
-
   |3ul
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   |5 ul
   |-
   |-
-
   |2mM sNTPs
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   |KOD plus neo
-
   |5ul
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   |1 ul
   |-
   |-
   |10xPCR buffer
   |10xPCR buffer
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PCR:60sec, 30 cycle 1h8m
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{|class="hokkaidou-table-pcr-time"
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|-
-
 
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|Number
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</p>
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|Degree
 +
|Second
 +
|-
 +
|1
 +
|94
 +
|120
 +
|-
 +
|2
 +
|98
 +
|10
 +
|-
 +
|3
 +
|68
 +
|60
 +
|-
 +
|4
 +
|4
 +
|HOLD
 +
|}
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Cycle:2~3 x 30
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Did not work well?
</div></div>
</div></div>
 +
<div class="hokkaidou-notebook-daily">
 +
===August 17th===
 +
<div class="hokkaidou-section">
 +
====Electrophoresis====
 +
We confirmed success of PCR of August 16th by electrophoresis.
 +
====Gel extraction====
 +
The plasmids done PCR were extracted from TBE gel, and then we got 50 ul DNA solutions.
 +
</div></div>
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Latest revision as of 01:24, 27 September 2012

Contents

August 16th

PCR

PCR of pGEM to get linear PhaA and PhaB using Laury primer and X-PhaA-up-F, S-PhaA-dw-R, X-PhaB-up-F, S-PhaB-dw-R.

pGEM(pGEM 1 ul + DW 9 ul) 1 ul
up primer(10 mM) 1 ul
down primer(10 mM) 1 ul
25 mM MgSO4 3 ul
2 mM sNTPs 5 ul
KOD plus neo 1 ul
10xPCR buffer 5 ul
DW 33 ul


Number Degree Second
1 94 120
2 98 10
3 68 60
4 4 HOLD

Cycle:2~3 x 30

Did not work well?

August 17th

Electrophoresis

We confirmed success of PCR of August 16th by electrophoresis.

Gel extraction

The plasmids done PCR were extracted from TBE gel, and then we got 50 ul DNA solutions.


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