?-easiest clean-up for sequencing

David L. Haviland, Ph.D. dhavilan at IMM2.IMM.UTH.TMC.EDU
Wed Feb 26 11:24:01 EST 1997


At 11:29 2/25/97 EDT, vilimf01 at mcrcr6.med.nyu.edu wrote:
>I knew this would come up sooner or later.  My answer is bound to piss
>off many of the suppliers out there.  We send our plasmids to a
>core facility for sequencing, and I have been playing around with
>different purification schemes to see which gives the best results. 
		** Edited **

Hi:

I'm not suprised by this in the least.  One question though, this IS for
automated sequencing, yes?

Last year I was doing transient transfections in to COS cells.  To optimize
"things", I had to determine which method was best suited for transfection
so I compared  the standard CsCl (Maniatis), PEG Alk-Lysis (BioTechn.
14:532), Wizzard minis, and Qiagen midis.  All rendered DNA that could be
visualized and digested.  

Without fail, the CsCl material was THE best for transfection followed by
the PEG Alk-lysis material.  A distant third was material prepared by the
Wizzard, and bringing up the rear was Qiagen.   The Promega B-gal vector
was prepared using all four methods.

This is what worked for me, "in my hands" of course.  But I think it does
go to show that "kits" don't often do what they claim.

David


=============================
 David L. Haviland, Ph.D.
 Asst. Prof. Immunology 
 University of Texas - Houston, H.S.C.
 Institute of Molecular Medicine  
 2121 W. Holcombe Blvd.  
 Houston, TX  77030 
 Internet:"dhavilan at imm2.imm.uth.tmc.edu" 
 Voice: 713.500.2413  FAX: 713.500.2424
 ------------------------------------------------------  
" Sometimes you're the windsheild, sometimes you're the bug."
=============================




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