Team:HokkaidoU Japan/Notebook/aggregation Week 9

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(27th colony pcr, incubation)
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==August 27th==
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<div>
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==Colony PCR==
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<p>
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Colony PCR to confirm that whether the pT7-RBS-Ag43-dT on pSB1C3 was successfully ligated or not. 
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{|class="hokkaidou-table-pcr-reagent"
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|-
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|DNA solution
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|4 ul
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|-
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|Kapa-Taq(Taq polymerase)
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|5 ul
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|-
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|Forward Primer(ag43-f4 primer)
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|0.5 ul
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|-
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|Reverse Primer(200bp down primer)
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|0.5 ul
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|-
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|Total
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|10 ul
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|}
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{|class="hokkaidou-table-pcr-time"
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|-
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|Number
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|Degree
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|Second
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|-
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|1
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|95
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|120
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|-
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|2
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|95
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|30
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|-
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|3
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|53.0
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|30
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|-
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|4
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|72
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|60
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|-
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|5
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|72
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|60
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|-
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|6
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|4
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|HOLD
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|}
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Cycle:2~4 x 35
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We used N1 (DW only) and N2 (Ag43-dT on pSB1AK3) as controls.
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Desired product is about 695bp.
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[[image:|thumb|Colony PCR result]]
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</p>
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==Incubation for mini-prep==
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<p>
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Incubation of pT7-RBS-Ag43-dT on pSB1C3 in LBC liquid medium.
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#Prepared 2 ml LBC into culture tubes.
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#Re-suspended 2 colonies (No.1 and No.2 respectively).
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#Incubated at 37C for 15 hrs.
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</p>
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</div></div>
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Revision as of 05:37, 28 August 2012

August 27th

Colony PCR

Colony PCR to confirm that whether the pT7-RBS-Ag43-dT on pSB1C3 was successfully ligated or not.

DNA solution 4 ul
Kapa-Taq(Taq polymerase) 5 ul
Forward Primer(ag43-f4 primer) 0.5 ul
Reverse Primer(200bp down primer) 0.5 ul
Total 10 ul


Number Degree Second
1 95 120
2 95 30
3 53.0 30
4 72 60
5 72 60
6 4 HOLD

Cycle:2~4 x 35

We used N1 (DW only) and N2 (Ag43-dT on pSB1AK3) as controls. Desired product is about 695bp.

[[image:|thumb|Colony PCR result]]



Incubation for mini-prep

Incubation of pT7-RBS-Ag43-dT on pSB1C3 in LBC liquid medium.

  1. Prepared 2 ml LBC into culture tubes.
  2. Re-suspended 2 colonies (No.1 and No.2 respectively).
  3. Incubated at 37C for 15 hrs.


Retrieved from "http://2012.igem.org/Team:HokkaidoU_Japan/Notebook/aggregation_Week_9"