In article <Pine.GSO.4.10.10012141136340.19433-
100000 at merhaba.cc.columbia.edu>,
"Gary G. Wilson" <ggw5 at columbia.edu> wrote:
> Can someone give me some idea of how to do an X-gal stain on a
bacterial
> plate? I've been given an old reference to Miller but don't have
time to
> wait to get it on interlibrary loan.
>> Do I add the X-gal to the agar?
Either directly to the freshly made agar (after autoclaving), or spread
on plate (more economical), the stuff is somewhat expensive
How much?
spread 40 ul of a 20 mg/ml solution (in DMF) per plate
(double-check please)
How long does it take before I
> see blue cells (typically)?
depends on strain, 8-16 h (ballpark), sometimes it helps to put plates
in fridge after ovn growth
Do I need to add anything else or just
> X-gal etc.
- when spreading dilute w/ LB
- may have to add IPTG (0.5 - 1 mM) in lacI+ background
>> A pointer to a contemporary reference
you have the original ref. (miller), another common ref.: Sambrook et
al., Molecular Cloning (probably somewhere around your department)
which detials this method would
> suffice if nobody wants to give a verbose reply.
>> Thanks
>> Gary G Wilson (attempting to learn some microbiology)
>>
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