Preparing an Adenoviral shRNA Clone - Revision history

Davebrid: /* Choosing a shRNA Sequence */ updated XbaI compatible overhang

2010-12-13T19:47:38Z

Choosing a shRNA Sequence: updated XbaI compatible overhang

← Older revision		Revision as of 19:47, 13 December 2010
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	==Choosing a shRNA Sequence==		==Choosing a shRNA Sequence==
	Use oligoengine to pick a sequence, for example		Use oligoengine to pick a sequence, for example
	<span style="color:#FF0000">~~GATC~~</span><span style="color:#00FFFF">CCC</span>GAATACCGCAATGCCTTAG<span style="color:#FFD700">TTCAAGAGA</span>CTAAGGCATTGCGGTATTC<span style="color:#00FFFF">TTTTTA</span>		<span style="color:#FF0000">CTAG</span><span style="color:#00FFFF">CCC</span>GAATACCGCAATGCCTTAG<span style="color:#FFD700">TTCAAGAGA</span>CTAAGGCATTGCGGTATTC<span style="color:#00FFFF">TTTTTA</span>

	Where:		Where:

2010-09-16T14:34:03Z

added categories

← Older revision		Revision as of 14:34, 16 September 2010
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	__NOTOC__		__NOTOC__
			[[Category:shRNA]][[Category:Adenovirus]][[Category:Transduction]][[Category:Knockdown]][[Category:Molecular Biology]][[Category:Cloning]]
	==Choosing a shRNA Sequence==		==Choosing a shRNA Sequence==
	Use oligoengine to pick a sequence, for example		Use oligoengine to pick a sequence, for example

2010-09-14T14:37:36Z

added protocol links

← Older revision		Revision as of 14:37, 14 September 2010
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	==Cloning Targetting Construct==		==Cloning Targetting Construct==
	*see http://www.oligoengine.com/products/psuper/documentation/protocols/pSUPER_protocol.pdf		*see [http://www.oligoengine.com/products/psuper/documentation/protocols/pSUPER_protocol.pdf pSuper] and [http://www.coloncancer.org/adeasy/He,%20AdEasy%20-%20Nature%20Protocols,%202007.pdf AdEasy] protocols
	===Digestion of pAdtrack-H1 Promoter (or pSuper)===		===Digestion of pAdtrack-H1 Promoter (or pSuper)===
	#If using a BglII/HindII do a sequential digestion		#If using a BglII/HindII do a sequential digestion

2010-08-26T13:40:58Z

no table of contents

← Older revision		Revision as of 13:40, 26 August 2010
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			__NOTOC__
	==Choosing a shRNA Sequence==		==Choosing a shRNA Sequence==
	Use oligoengine to pick a sequence, for example		Use oligoengine to pick a sequence, for example

2010-08-26T13:40:32Z

filled in protocol from http://www.oligoengine.com/products/psuper/documentation/protocols/pSUPER_protocol.pdf

@@ Line 4: / Line 4: @@
 Where:
-*<span style="color: #FF0000">XbaI compatible overhang</span>
+*<span style="color: #FF0000">BglII compatible overhang</span>
 *<span style="color: #00FFFF">Spacer</span>
 *<span style="color:#FFD700">Loop</span>
@@ Line 11: / Line 11: @@
 For the reverse primer choose the reverse complement, remove the XbaI overhang from the 3' end (the last GATC sequence) and add an AGTC sequence to the 5' end (for HindIII).
 Order primers through IDT-DNA

2010-08-23T18:21:15Z

← Older revision		Revision as of 18:21, 23 August 2010
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	*<span style="color: #00FFFF">Spacer</span>		*<span style="color: #00FFFF">Spacer</span>
	*<span style="color:#FFD700">Loop</span>		*<span style="color:#FFD700">Loop</span>
			*In black is the target shRNA sequence, both forward and then after the loop as a reverse complement.

	For the reverse primer choose the reverse complement, remove the XbaI overhang from the 3' end (the last GATC sequence) and add an AGTC sequence to the 5' end (for HindIII).		For the reverse primer choose the reverse complement, remove the XbaI overhang from the 3' end (the last GATC sequence) and add an AGTC sequence to the 5' end (for HindIII).
	Order primers through IDT-DNA		Order primers through IDT-DNA