Southern with small probe?

John Dixon jpcd0 at mole.bio.cam.ac.uk
Fri Mar 17 11:52:08 EST 1995


Hi, I just tried to do a southern with a 220 bp fragment. No bands at all.
The fragment is the supF minigene which I know is included in both LTR's
of some proviral insertions I am investigating.

Is 220 bp too small for some reason? - I have probed happily with 500bp
fragments (random priming in both cases with alpha 32P CTP)

If I used the whole 600bp LTR to probe would this work? (the supF replaces
central 240bps of +type LTR sequence)

Would this cross hybridise to any other retrovirus or retrovirus like
elements (my LTRs are MoMuLV and my genomic DNA is from 129 genetically
inbred mouse line, BTW)

Perhaps the supf minigene which codes for a tRNA molecule would be
expected to have stem loops and so on when single stranded and thus make a
crap probe (I hybed at 65%)

Any suggestions?

TIA

Johnny D

-- 
John Dixon                     Lab 44 (223) 334131
Wellcome/CRC Institute         Fax 44 (223) 334134
Dept Genetics
Cambridge University    
United Kingdom       e-m: jpcd0 at mole.bio.cam.ac.uk



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