PCR purification kit

ChenHA hzhen at freeuk.com
Mon May 22 06:08:10 EST 2000



Zhonglin Chai wrote:
> 
>     Depending on what you will use the purified pcr products for, you
> may have a varieties of choices there.
>     Personally I have been using Promega's Sepharcryl S400 matrix to
> purify or to get rid of primers of pcr products for sequencing. You just
> need to load the matrix to a spin column, and spin to dry the matrix,
> and then load pcr reactions (30uL or a bit more) to the surface of the
> matrix, and spin again for a few minutes to collect the DNA. Anything
> smaller than 400bp will retain in the column and the larger products
> flow through. You then just need 2-5uL directly for sequencing. They
> have also S300, namely able to get rid of DNA smaller than 300 bp. This
> was initially designed to get rid of small synthesized cDNA fragments
> for library construction, but it worked very well for pcr products
> purification for sequencing purpose in my hands.

That is probably not much use for my purpose as the pcr
products I use is typically in the range of 200-1000 bp. 
But thanks for the suggestion anyway, it is not one I have
used and it would be interesting to see if there is other MW
ranges.

>     If you need to clone the pcr products, you may need to get rid of
> the DNA polymerase and other junks as well. Either phenol/chloroform
> extraction or gel purification before restriction digestion seems to be
> necessary and worked equally well in my hands. Of course, after
> digestion you need to gel purify before doing ligation. 

The idea of using the kit is that it is fast and
convenient.  If it is necessary to phenol/chloroform or gel
purified, then I think there is not much advantage in using
kit.  

Cheers



I have been
> using Qiagen's Qiaquick Gel Extraction kit that works beautifully with
> satisfactory yield. BTW, elute the DNA in 10mM Tris solution provided in
> the kit. If you elute in water, either the yield may drop, or the DNA
> may migrate differently in agarose gel showing one or more additional
> bands with smaller molecular weights than expected, although the MW may
> become back to normal after adding tris or salt to the solution.
>
> --
> ZhongLin Chai, PhD
> __________________________________________________________
> Department of Pathology and Immunology
> Monash University Medical School
> Alfred Hospital
> Commercial Rd, Prahran, VIC 3181, AUSTRALIA
> Telephone: (61 3) 9903 0698 (lab)
>            (61 3) 9903 0696 (office)
> Mobile:    0413 58 1940 or International: +61 413 58 1940
> Fax:       (61 3) 9903 0731
> email:     zhonglin.chai at med.monash.edu.au
> ___________________________________________________________




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