Klaus.Matthaei at anu.edu.au Klaus.Matthaei at anu.edu.au
Wed Sep 23 18:18:36 EST 1992

>In regard to the thread on micropipette calibaration:
>Several people have mentioned that they do not bother recalibrating;
>they just clean carefully several times a year.  I have a horror story
>that has induced our lab into careful, frequent cleaning...
>For the past year, we have been having trouble with plasmid contamination
>on our genomic southern blots.  Sometimes one lane is affected, sometimes
>an entire gel.  I screamed and pleaded for people to be careful,threw out 
>all of the solutions in the lab several times, but the problem persisted.
>Finally I had the bright idea that our pipettepeople might be infested 
>with plasmid DNA.  After washing, the problem disappeared.  What had
>happened was that the person making our plasmid preps was using the P1000
>to resuspend pellets, and massively contaminating the pipette barrel.
>People doing genomic DNA preps were occassionally getting a little solution
>into the barrel and randomly contaminating their preps.  Now we wash
>them regularly, and keep separate pipettors for plasmid and genomic DNA
>(we already had a separate set for PCR).
>Andrew Cockburn


Ever wondered what those so called 'deletions' are when you subclone a
fragment and get vector only even if properly phosphatased?  Guaranteed
Gilson contamination. 
We routinely decontaminate our Pipetpeople (yes we have been using
non-sexist language for some time also) to overcome this problem.  We used
to depurinate the barrels with HCl followed by NaOH (0.1N and 0.4N resp.)
but now treat with sodium perchlorate (~5% for 5 min, see ref. that escapes
me at the moment in a Biotechniques issue suggested for PCR
decontamination) and have reduced, if not eliminated the problem (sloppy
people still exist). 

Hope this helps.

Cheers, Klaus

PS:  5X loading dye is also a great source of spurious bands in southerns
etc.from the same source and multi users.

Klaus Matthaei
Gene Targeting
The John Curtin School of Medical Research
The Australian National University
E-mail: Klaus.Matthaei at anu.edu.au

"If all else fails.  Read the instructions".

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