Microsatellite stutter bands

Lars P. Ryder ryderlp at pip.dknet.dk
Wed May 21 04:53:14 EST 1997


John Ladasky wrote:
> 
> Greetings,
> 
>         I'm doing a little microsatellite typing on the side of my regular
> project.  I'm dismayed to see several prominent secondary products.  My
> understanding is that these secondary peaks, called "stutter bands," arise
> from slipped-strand mispairing during PCR.  A colleague said that using *two*
> temperature thermal cycling (anneal and extend at the same temperature) is
> supposed to help reduce the stutter bands, but he didn't remember the refer-
> ence.  Can anyone vouch for this and/or remember the reference?  Do you have
> any other hot tips for cleaning up microsatellite PCR?  Thanks!


Two step PCR will minimize nontemplate nucleotide addition by Taq
(addition of extra A) - NOT stutter bands. Select tri- or
tetranucleotide repeats and you will see almost no stutter bands. The
ref is: Smith, Genome Res. 5:312-17 (1995).


Regards,

Flemming

-- 
Flemming Larsen 
Dept. of Clin. Immunol., sect. 7631
National University Hospital
Tagensvej 20
DK-2200 Copenhagen N
Denmark

E-mail: flar at biobase.dk
phone:  +45 35 45 78 48
fax:    +45 31 39 87 66



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