pea cDNA/B.napus Southern problem
bckraev at aeolus.ethz.ch
Thu Nov 10 04:50:12 EST 1994
Try to make a copy reconstruction experiment, that is apply 50 picograms, 25
picograms etc of the probe plasmid on your gel, run it, blot and hybridize.
If you see 25 ng plasmid band after overnight exposure under your conditions,
the method works. Your problem may be that the amount of your target in
genomic lanes does not match the sensitivity of your Southern. This is a
typical problem with large plant genomes. We had great success only with
probes >10E9/ug, using nylon filters and UV crosslinking.
Hope this helps, Sasha
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