Mwatson's 20 kbp PCR question.

KASRA RAMYAR cenobite at welchlink.welch.jhu.edu
Fri Aug 26 16:15:49 EST 1994


In article <aquilla.1126944584M at sadye.emba.uvm.edu>,
Tracy Aquilla <aquilla at salus.med.uvm.edu> wrote:
>In Article <1994Jul27.224302.5558 at news.cs.brandeis.edu>,
>story at binah.cc.brandeis.edu wrote:
>>Dear ?,
>>
>>One thing you should try for 20 kb PCR which is new on the market is PCR
>>extender additive from Stratagene. They will undoubtedly gove you a free sample
>>if you ask them. It's amazing stuff. 
>>
>>Also try the group bionet.molbio.methds-reagnts for ongoing discussion of the
>>topic of long range PCR.
>>
>>Cheers
>>
>>Craig Story
>>story at binah.cc.brandeis.edu
>>
>
>I just called, and they're not giving it away for free any more. Bummer!
>    Tracy
	
	Hmmmm, I'm truely curious as to the contents of the Extender, but 
if you are interested in doing long range PCR, then get KlenTaq1 from a 
company called:
		Ab Peptides, Inc.
		6272 Marmaduke Ave.
		St. Louis, MO 63139
		Tel (314) 644-1700
		Fax (314) 644-2100
This enzyme, according to their info sheet, is Taq poly, without some of 
the N-terminus.  So it loses its 5' exo domain.  When they originally 
marketed this enzyme, they were bundling it with Pfu from Stratagene, 
mixing it in a 1:6 (Pfu:KlenTaq1) ratio (by unit).  Then I guess they got 
sued for selling Stratagene enzymes and they stopped selling the mix (I 
think they were calling it KlenTaqLA).  The reason for mixing the two is 
that KlenTaq1 cannot proofread, yet Pfu has a much lower processivity and 
falls off, so working in tandem ...
	Someone in my lab used the stuff to PCR genomic inserts out of 
EMBL3.  The PCR was very specific and yielded a 12.5-13 Kb band.  Not 
bad, huh!  So I wonder if Taq Extender (tm) is actually a thermophillic 
poly that is more processive than Pfu. 
	Hope this helps.

							Kas
 




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