What does A, C rich sequence mean?
cyk10 at cus.cam.ac.uk
Fri Jan 28 08:22:39 EST 1994
Why don't you try using exoIII deletions and continue to (successfully)
use the vector primer to sequence? A method is given in Maniatis (orig.
Henikoff, of course). You'd probably want to do the digests at 30 deg C
(or even cooler) and take 1/2 min timepoints instead of 1 min (seeing that
you only have 600 b.p. of sequence). In my hands the digestion rate at
30 deg C is about 250 bp a minute. Wonderful method, the reaction takes
quarter of a day (after setting up and making solutions etc), you do the
transformations and hey presto! get colonies the next day. pick colonies,
do overnights, then minipreps/single strand rescues and you're ready to
sequence. Much, _much_ faster than walking...
Hope this helps!
Gong Xi Xin Nian!
Chong-Yee Khoo E-mail: cyk10 at cus.cam.ac.uk
Wellcome/CRC Institute Talk: cyk10 at cy-mac.welc.cam.ac.uk
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