expression in minimal media - (Apr/03/2009 )

I've done some more work on this problem- and now my expression works!

It was the trace elements mix (1000x)! The mix tends to oxidise, which makes some metals fall out of solution and become unavailable for uptake by the bacteria. I made up the mix again- and my expression in M9 works. The trace elements are important for the TCA cycle and other metal dependant cell processes. Thank heavens.

Trace elements mix should NOT be dark brown or rusty colour, but a golden/dark yellow colour. Don't keep the mix stored for too long.

Microbiology- part science, part witchcraft!

Cheers
LB

Luria Bertani on Wed Jun 10 23:19:54 2009 said:

Hi!
after I sterile filter my 1000x trace metal, the color is dark yellow too! I'm trying to figure out which metal fall out of my solution, because the concentration might be off after some metals crash out.
my 1000x trace metal recipe is:
a. 50mM FeSo4.7H2O (no problem dissolving it)
b. 10mM MnCl2.4H2O (no problem dissolving it)
c. 2mM CoCl2.6H2O (no problem dissolving it)
d. 10mM ZnSo4.7H2O (no problem dissolving it)
e. 2mM NiSO4.6H2O (no problem dissolving it)
f. 2mM (NH4)6Mo7O27.4H2O (no problem dissolving it) but color turned from yellow to RED PURPLE!!!
g. 20mM CaCl2 (no problem dissolving it)
h. 2mM CuCl2 (no problem dissolving it)
i. 2mM H3BO3 (slight issue with solubility, can see cloudiness) maybe is h or g, not sure

any pointers on solubility issues and oxidizing problems would be greatly appreciated!
THANKS

adjust the pH (to 7?)

mdfenko on Fri Apr 1 18:48:02 2011 said:

Hi,
Thanks for replying. I don't think it's pH problem, dilute HCl actually helps to keep Iron in solution.

I'm thinking about removing the (NH4)6Mo7O27.4H2O from my trace metal, the dark color might be metal oxides. Any ideas? I really don't want formation of metal oxides.

Thanks in advance!
ILJ

what IPTG concentration do you use for induction?

Regards,
p