betaine in LA PCR

Wayne M. Barnes wayne at BARNES1.WUSTL.EDU
Thu Apr 17 23:30:29 EST 1997


Dear Brad and Paul,

     The reference below is to using betaine in PCR.  We find it
enhances LA PCR up to 7 kb.  We worry about a negative effect on fidelity,
but we don't have data on this yet.  We leave out the DMSO from the reference
below, except for 1% sometimes.  

     Add 1.3 M final betaine, from a 4 M or 5 M stock.  

     Take one or two degrees off the melting temp of your PCR cycles.

     We find that even ordinary 5 kb PCR is enhanced, not just the high 
GC targets targeted by the reference below.


[2] Baskaran, N., Kandpal, R.P., Bhargava, A.K., Glynn, M.W., Bale, A., &
Weissman, S.M. (1996) Uniform amplification of a mixture of
deoxyyribonucleic acids with varying GC content, Genome Research 6:633-638.


Wayne M. Barnes, Ph.D.                        wayne at barnes1.wustl.edu
Biochemistry Dept. 8231                   or  barnes at biodec.wustl.edu
Washington Univ. Medical School               314.362.3351  fax 7183
660 South Euclid Ave., St. Louis, MO 63110
           http://mbb.wustl.edu/~barnes/

Dear Wayne,

I was interested in your post and was wondering if you had a reference
or a protocol for the use of betaine in cycle sequencing.  Also, is
betaine generally advantageous to regular or long pcr?

Thanks, 
Brad Turner

**************************************************************
		    Bradley Turner
	        Beth Israel Deaconess
                    Medical Center

Harvard Medical School		617-667-1215 phone
Division of Gastroenterology	617-667-2767 fax
Room Dana 536			bsturner at mbcrr.harvard.edu
330 Brookline Avenue		bturner at bidmc.harvard.edu
Boston, MA 02215, USA		turner at sprcore.bidmc.harvard.edu
**************************************************************



Dear Wayne:

I read on the methods group that you are recommending adding
1.3-1.5 M betaine for LA PCR and cycle sequencing. Can you
give me any more information as to what you think is going on
and any references for betaine in PCR?

Thanks very much.

--
Paul N. Hengen, Ph.D.
National Cancer Institute
mailto:pnh at ncifcrf.gov
http://www-lmmb.ncifcrf.gov/~pnh/




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