[Q] detecting a mutation site on a BAC clone
Michael J. Prigge
prigge at darkwing.uoregon.edu
Mon Dec 2 23:55:03 EST 1996
In article <32A3901F.17FE at bric.postech.ac.kr>, hgn at WWW.BRIC.POSTECH.AC.KR
(Hong- Gil Nam) wrote:
> Dear netters:
> I am trying to clone a gene for gi mutataion in Arabidopsis. We have
> identified a BAC clone that are very likely to encompass the mutation.
> We wish to find where the mutation site is on the BAC clone to get a
> clone for funtional complementation by transformation. The BAC clone is
> from wild type. We have three independent alleles of mutation, two of
> which are by X-ray and gamma-ray mutations.
I would label up the BAC DNA and hybridize it to WT plus the mutants'
DNA restricted with various enzymes. With the Arabidopsis BACs I've
looked at, HindIII, for example, gives a spread in fragment sizes
from 300 bp to 6 kb. One should be able to detect a deletion of <100
bp if one exists in your mutants.
Even if the odds are not in your favor, its an easy experiment.
Good Luck. Let me know how it comes out (if you try) as I may be in a
similar situation soon.
Michael Prigge prigge at darkwing.uoregon.edu
Institute of Molecular Biology University of Oregon
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