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Revision as of 14:39, 17 September 2007 (view source) Eismoustique (Talk | contribs) ← Older edit		Latest revision as of 18:05, 7 October 2007 (view source) Nicolas C. (Talk | contribs)
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		+	[[Paris/September 16|yesterday]] -- [[Paris/September 18|tomorrow]] <br>
	== DapA Wanner deletion: pKD13 - DapA homologies PCR ==		== DapA Wanner deletion: pKD13 - DapA homologies PCR ==
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	|width=1%|		|width=1%|
	|- style="background: #cccccc;"		|- style="background: #cccccc;"
-	| style="background: #FF99FF;" |D56	+	| style="background: #ccffcc;" |D56
	|araC/pBad>>RBS-Cre BBinsert		|araC/pBad>>RBS-Cre BBinsert
	|L26.1 araC/pbad>>RBS-Cre in <bbpart>J61002</bbpart>		|L26.1 araC/pbad>>RBS-Cre in <bbpart>J61002</bbpart>
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	|		|
	|- style="background: #cccccc;"		|- style="background: #cccccc;"
-	| style="background: #ccffcc;" |D72	+	| style="background: #FF99FF;" |D72
	|<bbpart>pSB2K3</bbpart> BBvector		|<bbpart>pSB2K3</bbpart> BBvector
	|MP16 <bbpart>pSB2K3</bbpart>::attP		|MP16 <bbpart>pSB2K3</bbpart>::attP
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	|		|
	|- style="background: #cccccc;"		|- style="background: #cccccc;"
-	| style="background: #ccffcc;" |D73	+	| style="background: #FF99FF;" |D73
-	|<bbpart>pSB2K3</bbpart> BBvector	+	|<bbpart>pSB2K3</bbpart> ES open vector
	|MP16 <bbpart>pSB2K3</bbpart>::attP		|MP16 <bbpart>pSB2K3</bbpart>::attP
	|EcoRI		|EcoRI

---

Latest revision as of 18:05, 7 October 2007

yesterday -- tomorrow

DapA Wanner deletion: pKD13 - DapA homologies PCR

• Material:
  

- pKD13: 5 µL (bacteria)
- Tampon GC: 10µL
- dNTP: 1µL (final 200µmol each)
- Taq: 0.5µL
- Water: 32.5µL
- Primer1: 33: 2.5µL (final 0.5µM)
- Primer2: 34: 2.5µL (final 0.5µM)

File:WannerDapA09172007.jpg

• PCR Program:
  

35 cycles:
- 98°C: 5min
- 98°C: 45s (X35)
- 69°C: 30s (X35)
- 72°C: 1min (X35)
- 72°C: 10 min
- 4°C: inf

• Quantity: ng/µL
  
• Expected size: 1403 kb

Digestions

Digestion Products
Number	Product Name	Matrix Name	Enzyme 1	Enzyme 2	Size	Description
D56	araC/pBad>>RBS-Cre BBinsert	L26.1 araC/pbad>>RBS-Cre in J61002	EcoRI	Pst1		araC/pBad>>RBS-Cre BBinsert (for transfer to the low-copy plasmid vector)
D66	KanR BBa_J61025 FV	(MP12.1, MP12.2) BBa_P1003 in pSB1A2	EcoR	XbaI		KanR BBa_J61025 (for forward introduction of a casette for Wanner K insertion)
D71	araC/pBad>>lox71-gfptripart FI	L35.1 araC/pBad>>lox71-gfptripart BBa_J61002	EcoRI	SpeI		araC/pBad>>lox71-gfptripart for use as a FI
D72	pSB2K3 BBvector	MP16 pSB2K3::attP	EcoRI	PstI		Low copy number plasmid, BB vector
D73	pSB2K3 ES open vector	MP16 pSB2K3::attP	EcoRI	SpeI		Comparison of digestion efficiency between Pst and Spe in NEB2
D75	AraC-pBAD-RBS-CRE FI	L26.1 AraC-pBAD-RBS-CRE	EcoRI	SpeI		FI, Comparison of digestion efficiency between Pst and Spe in NEB2 (see D56)
D76	DGAT BB	P14 DGAT	EcoRI	PstI		DGAT