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hygromycin and arabidopsis

hygromycin and arabidopsis - Protocols and Methods Forum

hygromycin and arabidopsis - Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.


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Old 11-01-2007, 04:34 AM
Michael J. Prigge
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Default hygromycin and arabidopsis



I would guess that the false positive could be due to one of two
reasons. You could blast your primers against the Arabidopsis genome
to see if, by chance, there is 500 bp region flanked by sequences
similar to one or both of your primers. You could also check the
sequence or restriction pattern to confirm that the 500 bp fragment
is what you expect. More likely, one or more of your pipettors is
contaminated (from pipetting the plasmid or PCR products). Did you
do a no-DNA control? If you also see the product, then you'll
probably have to replace your reagents and clean your pipettors
(especially inside the shaft) or borrow someone else's for setting up
the reactions. (UV treatment has been shown to reduce contamination,
too.) Pipettor contamination can happen surprisingly easily,
especially if you pipet up and down to mix in dye or to rinse tips
between loading samples on the gel. I routinely use a different
pipettor for loading PCR products on gels.
Good Luck,
Mike

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