Team:HokkaidoU Japan/Notebook/August19

From 2010.igem.org

(Difference between revisions)
 
(10 intermediate revisions not shown)
Line 1: Line 1:
-
{{Template:HokkaidoU_Japan}}
+
{{Template:HokkaidoU_Japan}}<div class="linkbar"><div class="prev">[[Team:HokkaidoU_Japan/Notebook/August18|August 18]]</div>[[Team:HokkaidoU_Japan/Notebook|Notebook]]<div class="next">[[Team:HokkaidoU_Japan/Notebook/August23|August 23]]</div></div>
 +
 
=Ligation=
=Ligation=
-
VectorにHeat shock promotorとRBSを組み込む
+
Ligated Vector with Heat shock promotor and RBS
-
{|style="text-align:center;"
+
{|border="1px" class="protocol"
|-
|-
-
|style="border-bottom:1px solid #000;"|パーツ
+
!Parts
-
|style="border-bottom:1px solid #000;"|Markerと比べて (ng/uL)
+
!By comparison to marker
-
|style="border-bottom:1px solid #000;"|size (bp)
+
!size (bp)
-
|style="border-bottom:1px solid #000;"|必要 (ng)
+
!(ng) required
-
|style="border-bottom:1px solid #000;"|使用 (uL)
+
!(uL) used
|-
|-
|Vector
|Vector
|5 ng/uL
|5 ng/uL
-
|2000 bp
+
|2996 bp
|10 ng
|10 ng
|2 uL
|2 uL
Line 28: Line 29:
|1 uL
|1 uL
|-
|-
-
|style="border-top:1px solid #000;" colspan="4"|'''Total'''
+
|style="border-top:1px solid #996;" colspan="4"|'''Total'''
-
|style="border-top:1px solid #000;"|'''3.3 uL'''
+
|style="border-top:1px solid #996;"|'''3.3 uL'''
|}
|}
-
その後は，先日のLigation & Transformationと同じく．
+
Ligation and [[Team:HokkaidoU_Japan/Protocols|transformation]] follows same protocol as previously
-
 
+
-
=competencyを求める=
+
=Competency Check=
-
# 0.35 ug/uLのpUC119 DNAを1 uL 0.5 mLチューブにとり，2.5 uLのDWを加えた (0.1 ug/uL)
+
# 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)
-
# 999 uLのDWを1.5 mLチューブにとり，1 uLの[1]を加えた (0.1 ng/uL)
+
# Took new 1.5 mL tube and added 999 uL of ADW (0.1 ng/uL) to it plus 1 uL of [1]
-
# 99 uLのDWを1.5 mLチューブにとり，1 uLの[2]を加えた (1 pg/uL)
+
# Took new 1.5 mL tube and added 99 uL of ADW to it plus 1 uL of [2]
-
1 pg/uLのpUC119 DNAを「青α」と「赤α」のコンピテントセルに，それぞれ1 uL, 10 uL, 100 uLずつ加え，Transformationさせた
+
*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 [[Team:HokkaidoU_Japan/Protocols|Transformation]] between Vector，Transformation=
-
=VectorだけでLigation，Transformation=
+
Because there weren't any colonies the previous day we suspected that there might be something wrong with vectors<br>
-
前日のLigationでコロニーができなかったので，ベクターに問題があるかもしれない．<br>
+
* Mixed 2 uL of pSB1C3, 2 uL of ligation solution and 0.4 uL of T4 ligase
-
* pSB1C3 2 uLにligation solution 2 uLとT4 ligase 0.4 uLを加えた
+
* Transformed
-
* Transformation
+

Latest revision as of 08:29, 27 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
Retrieved from "http://2010.igem.org/Team:HokkaidoU_Japan/Notebook/August19"