Revision as of 17:33, 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

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)
Took new 1.5 mL tube and added 999 uL of ADW (0.1 ng/uL) to it plus 1 uL of [1]
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

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．

@@ Line 34: / Line 34: @@
 Ligation and 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]
-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
 =VectorだけでLigation，Transformation=