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	#pUC19-FliC vector とinsertオリゴをライゲーション.		#pUC19-FliC vector とinsertオリゴをライゲーション.
	[混ぜ表]		[混ぜ表]
-	#	+	*Template1μL
		+	*Primer(FとR)5μL
		+	*10×buffer10μL
		+	*dNT

---

Revision as of 03:08, 27 October 2007

• pUC19 vectorにFliCをコードする遺伝子を挿入→pUC19-FliC

1. design primer(エキサイトPCRのため)

[PCR表]FliCの可変領域のうち、フィラメントの外側部分から５か所を選び、プライマーを設計(207,228,240,252,267)

1. エキサイトPCR→pUC19-FliC-vector
2. design insert(ヒスチジンタグ)

• six histidine loop peptide ("His-loop")
• -Ser-Ser-His-His-His-His-His-His-Ser-Ser,encoded by the oligo 5'-TCT-TCT-CAT-CAT-CAT-CAT-CAT-CAT-TCC-TCC-3' and anti oligo 5'-GGA-GGA-ATG-ATG-ATG-ATG-ATG-ATG-AGA-AGA-3'

1. pUC19-FliC vector とinsertオリゴをライゲーション.

[混ぜ表]

• Template1μL
• Primer(FとR)5μL
• 10×buffer10μL
• dNT