Ivonne

You sterilise everything by autoclaving, but the glucose in a separate bottle, and the phosphate buffers in a separate bottle. When combining them all together, they don’t autoclave so well, but separately they’re fine.

Good luck

How much of the PDM podwer should be added to 1 liter of diH2O?

CK

As I understand it, it allows greater cell densities to be reached, and the buffering and salts allow for greater plasmid stability, the glucose allows for quicker cell growth. Apart from that, I don’t think there is much more,

Thanks Sudheer, you pimp that media.

Cheers

Looks promising based on the feedback here. The lazier option: inoculating your LB overnight 3 hours earlier than usual?

Sure you can filter glucose, but as long as it is autoclaved with yeast extract or another amino acid source, there will be no caramel, I’ve done it plenty times and never had a hassle.

I tried this medium and got a whopping 6.5x increase in plasmid DNA production compared to LB! Highly recommended!!

Simple idea behind PDM is that the chemicals in the media “react” in the cell to form complex biostructures. They calculated the optimal concentration of each ingredient that would give you a high biomass and lots of plasmid. When you are isolating plasmid, you want your cells to divide like crazy without worrying about spending their energy on transcription or translation, obviously aside from the required amount to sustain cell growth. Hence, PDM would probably suck for protein isolation, although LB or TB can be used for this purpose because of the fact that they are not optimized for constant cell division and low protein synthesis.

About the autoclaving: Glucose will not brown unless heated with amino acids (called a Maillard reaction) as Liam said. Phosphates should be autoclaved separately as usual.

With this medium, I have successfully isolated 60-70 mg pure high-copy plasmid from 1L cultures (no kits involved).

I made a 10x solution of KH2PO4 + Na2HPO4, 10x Glucose, and a 1.2x of the rest. I autoclaved them separately, but when I mixed together something salted out and won’t dissolve, even upon boiling. I’ll repeat to once more to make sure I didn’t screw up the weighing, but this should work right?

Hi guys

Yes, sometimes something does not dissolve or precipitates out, and I am not sure which component it is. I suspect it is the phosphate salts, and so I generally cool everything to room temperature or lower before combining, and keep a stirrer in the bottle to make sure everything mixes properly. It seems to happen more often with agar plates, so I cool everything in a 55oC waterbath before combining, although the buffer and glucose is alot cooler than that by the time it is added.