Advse needed on DNA strand-separation

Karen Browning kbrowning at MAIL.UTEXAS.EDU
Fri Nov 11 11:28:08 EST 1994


>>Hi, folks. I would like to get some advise on DNA strand separation. I have 1 
>>mg of dsDNA of 150 nt long obtained by PCR and would like to separate each 
>>strand into ssDNA. Due to the large scale, the avidin-agrose method didn't 
work
>>well (here one strand of the dsDNA contains a bitin at terminus). I intend to
>>separate the dsDNA by denaturing PAGE (sequencing gel)because upon
>>denaturation, one strand has the extra biotin part which would slow down its
>>migration.
>>Any suggstions?
>>
>>Thanks
>>
>>Baohua
>>Ohio State University
>I can't quite read your article, but if your ds PCR fragment was produced
>with one biotinylated primer, then it is very simple to use Dynals 
>streptavidin coated magnetic beads to separate the two strands for sequencing
>G Jenkins
>

Try an alkaline agarose gel for strand separation.  That is how it was done 
in the dark ages of chemical sequences.




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