384-well bacterial growth

marc crepeau mwcrepeau at hawaii.rr.com
Wed Jun 28 17:40:26 EST 2006


I'm trying to do some library construction/screening using a robot to
pick colonies and print filters.  The initial picking into 384-well
shallow plates works okay and I get decent overnight growth in 50 ul
aliquots of LB, but I'm having problems making replica plates.  I use a
384-pin replicating tool to inoculate from the mother plate to a
daughter plate containing 50 ul of fresh LB.  After overnight growth
I'm getting very patchy results.  Only a small minority of the wells
show the obvious turbidity observed in the wells of the mother plate.
The remaining wells are not very turbid at all, to the point where most
are difficult to distinguish by eye from an uninoculated well.  Using
the replicator tool to print to agar from the mother plate yields
colonies, so I think I am probably managing to inoculate the liquid
cultures with at least a small amount of inoculant.  But they aren't
growing well.  This is in Nunc plates, catalog number 62409-604.  The
vector is pZErO-2 and the host strain is DH5a.

I wonder if anyone has any tips for growing E. coli in 384-well shallow
plates?  Googling the issue I find that many people use special plate
incubators with forced oxygenation, but I don't have access to one.
What else might work?



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