sequencing riddle...

Julian Parkhill J.Parkhill at bham.ac.uk
Tue Apr 8 12:14:09 EST 1997


In article <199704042300.QAA145216 at nestor.NMSU.Edu>, hroychow at NMSU.EDU
(Hiranya Roychowdhury) wrote:

> Hello All,
>         We have come across an interesting brain teaser. One of our
colleagues has amplified a piece of DNA from a plant using a single primer
(i.e. amplified region bordered by inverted repeats...). Now the teaser
(may not be one for some of you): How can this piece of DNA be sequenced
WITHOUT having to CLONE it? In other words, the investigator wants to use
the same primers to sequence the pcr products directly after purifying
them!

As another poster suggested, first find an enzyme that cuts the fragment once.

Then gel-purify the two bands and sequence each separately with the single
primer.

If the fragments are less than about 500 bp, you may find that it is
better to purify them from acrylamide.

To get the full sequence, you may have to design internal primers to read
back to the ends.

Julian.

-- 
          Julian Parkhill   -   J.Parkhill at bham.ac.uk
   CRC Laboratories, Inst. of Cancer Studies, Medical School,
 University of Birmingham, Edgbaston, Birmingham, B15 2TJ, U.K.
Disclaimer: If I thought anybody gave a damn about my opinions....



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