Miniprep/Kit-free high-throughput protocol - Revision history

Vaishnavi Ananth: /* BioStream version */

Vaishnavi Ananth — Thu, 19 Nov 2009 09:48:49 GMT

BioStream version

←Older revision		Revision as of 09:48, 19 November 2009
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	[[Category:Escherichia coli]]		[[Category:Escherichia coli]]

-	==~~BioStream~~ version==	+	==BioCoder version==
-	Following is the Miniprep/Kit-free high throughput protocol in ~~BioStream~~, a high-level programming language for expressing biology protocols. What you see here is the auto-generated text ouput of the protocol that was coded up in ~~BioStream~~ (see Source code). More information about ~~BioStream~~ can be found on my home page. Feel free to mail me your comments/ suggestions.[[User:Vaishnavi Ananth\|Vaishnavi]]	+	Following is the Miniprep/Kit-free high throughput protocol in BioCoder, a high-level programming language for expressing biology protocols. What you see here is the auto-generated text ouput of the protocol that was coded up in BioCoder (see Source code). More information about BioCoder can be found on my home page. Feel free to mail me your comments/ suggestions.[[User:Vaishnavi Ananth\|Vaishnavi]]
	====Text Output====		====Text Output====
	[[Miniprep - Kit-free high throughput protocol]]		[[Miniprep - Kit-free high throughput protocol]]
	====Source Code====		====Source Code====
	[[Miniprep - Kit-free high throughput protocol - source code]]		[[Miniprep - Kit-free high throughput protocol - source code]]

Vaishnavi Ananth at 05:04, 23 September 2009

Vaishnavi Ananth — Wed, 23 Sep 2009 05:04:47 GMT

←Older revision		Revision as of 05:04, 23 September 2009
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	[[Category:In vitro]]		[[Category:In vitro]]
	[[Category:Escherichia coli]]		[[Category:Escherichia coli]]
		+
		+	==BioStream version==
		+	Following is the Miniprep/Kit-free high throughput protocol in BioStream, a high-level programming language for expressing biology protocols. What you see here is the auto-generated text ouput of the protocol that was coded up in BioStream (see Source code). More information about BioStream can be found on my home page. Feel free to mail me your comments/ suggestions.[[User:Vaishnavi Ananth\|Vaishnavi]]
		+	====Text Output====
		+	[[Miniprep - Kit-free high throughput protocol]]
		+	====Source Code====
		+	[[Miniprep - Kit-free high throughput protocol - source code]]

Torsten Waldminghaus at 17:06, 23 February 2009

Torsten Waldminghaus — Mon, 23 Feb 2009 17:06:47 GMT

←Older revision		Revision as of 17:06, 23 February 2009
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		+	{{back to protocols}}
	==Background==		==Background==
	This protocol is adapted from "Molecular Cloning: A Laboratory Manual", Second Edition, Sambrook, Fritsch, and Maniatis. It is a quick, inexpensive way to purify large numbers of plasmids (I used to routinely do 80 at a time.--[[User:Kathmc\|Kathleen]]) and yields DNA that is clean enough for sequencing or for use as a PCR template.		This protocol is adapted from "Molecular Cloning: A Laboratory Manual", Second Edition, Sambrook, Fritsch, and Maniatis. It is a quick, inexpensive way to purify large numbers of plasmids (I used to routinely do 80 at a time.--[[User:Kathmc\|Kathleen]]) and yields DNA that is clean enough for sequencing or for use as a PCR template.

Reshma P. Shetty at 01:13, 16 May 2007

Reshma P. Shetty — Wed, 16 May 2007 01:13:56 GMT

←Older revision		Revision as of 01:13, 16 May 2007
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	[[Category:DNA]]		[[Category:DNA]]
	[[Category:In vitro]]		[[Category:In vitro]]
		+	[[Category:Escherichia coli]]

Madhadron at 22:41, 24 March 2007

Madhadron — Sat, 24 Mar 2007 22:41:16 GMT

←Older revision		Revision as of 22:41, 24 March 2007
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	#Spin for 30 min in a tabletop centrifuge at maximum speed at 4 ˚C.		#Spin for 30 min in a tabletop centrifuge at maximum speed at 4 ˚C.
	#Remove pellet from each tube with a toothpick. The cellular debris should stick well to the toothpick. Try to insert and remove the toothpick from the center of the tube so you don't get any cellular debris on the sides of the tube.		#Remove pellet from each tube with a toothpick. The cellular debris should stick well to the toothpick. Try to insert and remove the toothpick from the center of the tube so you don't get any cellular debris on the sides of the tube.
-	#Add 300 μL ice cold isopropanol to precipitate the DNA.	+	#Add 300 μL ice cold isopropanol to precipitate the DNA (or 300μL of 2:1 isopropanol:ammonium acetate, mixed just before you use it. See [http://www.madsci.org/posts/archives/2000-07/965055056.Mb.r.html this discussion] of precipitating nucleic acids.)
	#Spin for 10 min in a tabletop centrifuge at maximum speed at 4 ˚C.		#Spin for 10 min in a tabletop centrifuge at maximum speed at 4 ˚C.
	#Remove supernatent.		#Remove supernatent.

Jenny T Nguyen at 18:50, 10 July 2006

Jenny T Nguyen — Mon, 10 Jul 2006 18:50:43 GMT

←Older revision		Revision as of 18:50, 10 July 2006
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	10 mM Tris-HCl, pH 8<br>		10 mM Tris-HCl, pH 8<br>
	1 mM EDTA		1 mM EDTA
		+
		+	[[Category:Protocol]]
		+	[[Category:DNA]]
		+	[[Category:In vitro]]

Kathmc at 18:07, 11 January 2006

Kathmc — Wed, 11 Jan 2006 18:07:09 GMT

New page

==Background==
This protocol is adapted from "Molecular Cloning: A Laboratory Manual", Second Edition, Sambrook, Fritsch, and Maniatis. It is a quick, inexpensive way to purify large numbers of plasmids (I used to routinely do 80 at a time.--[[User:Kathmc|Kathleen]]) and yields DNA that is clean enough for sequencing or for use as a PCR template.

==Protocol==
#Transfer 1.5 mL of an overnight culture containing your plasmid to an eppendorf tube and spin at 5000 rpm for 5 min in a tabletop centrifuge to pellet the cells.
#Remove and discard the supernatent.
#Add 300 μL STET buffer, and resuspend cells by vortexing.
#Add 10 μL lysozyme (10 mg/mL), vortex, and submerse in boiling water for 40 sec.
#Spin for 30 min in a tabletop centrifuge at maximum speed at 4 ˚C.
#Remove pellet from each tube with a toothpick. The cellular debris should stick well to the toothpick. Try to insert and remove the toothpick from the center of the tube so you don't get any cellular debris on the sides of the tube.
#Add 300 μL ice cold isopropanol to precipitate the DNA.
#Spin for 10 min in a tabletop centrifuge at maximum speed at 4 ˚C.
#Remove supernatent.
#Add 200 μL ice cold 80% ethanol to wash pellet and spin for 5 min in a tabletop centrifuge at maximum speed at 4 ˚C.
#Remove supernatent.
#Dry pellet (air dry at room temperature or 37 ˚C or dry in a speedvac).
#Rehydrate in 50 μL TE.

==Buffers==
===STET===
8% sucrose 
50 mM Tris-HCl, pH 8 
0.5% Triton X-100 
50 mM EDTA 

===TE===
10 mM Tris-HCl, pH 8 
1 mM EDTA