Team:HokkaidoU Japan/Notebook/August19

From 2010.igem.org

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=Ligation=
=Ligation=
Ligated Vector with Heat shock promotor and RBS
Ligated Vector with Heat shock promotor and RBS
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{|class="protocol"
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{|border="1px" class="protocol"
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!Parts
!Parts
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|style="border-top:1px solid #996;"|'''3.3 uL'''
|style="border-top:1px solid #996;"|'''3.3 uL'''
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Ligation and transformation follows same protocol as previously
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Ligation and [[Team:HokkaidoU_Japan/Protocols|transformation]] follows same protocol as previously
=Competency Check=
=Competency Check=
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*Transformed
*Transformed
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=Ligation and Transformation between Vector，Transformation=
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=Ligation and [[Team:HokkaidoU_Japan/Protocols|Transformation]] between Vector，Transformation=
Because there weren`t any colonies the previous day we suspected that there might be something wrong with vectors<br>
Because there weren`t any colonies the previous day we suspected that there might be something wrong with vectors<br>
* Mixed 2 uL of pSB1C3, 2 uL of ligation solution and 0.4 uL of T4 ligase
* Mixed 2 uL of pSB1C3, 2 uL of ligation solution and 0.4 uL of T4 ligase
* Transformed
* Transformed

Revision as of 08:35, 2 October 2010

Notebook

Ligation

Ligated Vector with Heat shock promotor and RBS

Parts By comparison to marker size (bp) (ng) required (uL) used
Vector 5 ng/uL 2996 bp 10 ng 2 uL
Heat shock promotor 25 ng/uL 800 bp 8 ng 0.3 uL
RBS 1 ng/uL 50 bp 1 ng 1 uL
Total 3.3 uL

Ligation and transformation follows same protocol as previously

Competency Check

  1. Transfered 1 uL of pUC119 DNA (0.35 ug/uL) into 0.5 mL tube and added 2.5 uL of ADW (0.1 ug/uL)
  2. Took new 1.5 mL tube and added 999 uL of ADW (0.1 ng/uL) to it plus 1 uL of [1]
  3. Took new 1.5 mL tube and added 99 uL of ADW to it plus 1 uL of [2]
  • Added pUC119 DNA (1 pg/uL) to each DH5alpha tube of 50ul and 30ul also varying the amount of pUC119 DNA:1 uL, 10 uL, 100 uL
  • Transformed

Ligation and Transformation between Vector，Transformation

Because there weren`t any colonies the previous day we suspected that there might be something wrong with vectors

  • Mixed 2 uL of pSB1C3, 2 uL of ligation solution and 0.4 uL of T4 ligase
  • Transformed
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