Team:HokkaidoU Japan/Notebook/August19

From 2010.igem.org

(Difference between revisions)
(Ligation)
(Ligation)
Line 6: Line 6:
|-
|-
!パーツ
!パーツ
-
!Markerと比べて (ng/uL)
+
!By comparison to marker
!size (bp)
!size (bp)
-
!必要 (ng)
+
!(ng) required
-
!使用 (uL)
+
!(uL) used
|-
|-
|Vector
|Vector
Line 32: Line 32:
|style="border-top:1px solid #996;"|'''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 transformation follows same protocol as previously
=competencyを求める=
=competencyを求める=

Revision as of 16:50, 21 September 2010

Ligation

VectorにHeat shock promotorとRBSを組み込む

パーツ 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を求める

  1. 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)
  2. 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)
  3. 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)

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


VectorだけでLigation,Transformation

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.

  • pSB1C3 2 uLにligation solution 2 uLとT4 ligase 0.4 uLを加えた
  • Transformation