Dear Thomas,

we are using the same medium you use (HL5 without glucose, Formedium) and
add atoclaved Glucose seperately before use. At the moment we are using
several plasmids from the dicty stock center (pDXA-GFP2, pTX, pDM
series)that all have G418 resistance cassettes. We use G418 sulfate from
Calbiochem (Cat#345810) as a pouder. We solve it in water and sterile
filtrate and use it at a concentration of 10 µg/ml. It works very well. I
think I have all these information from you?! So if I can help by sending
aliquots of our DNA or G418 or HL5 so you can test in your lab, let me
know.

Greetings from Kiel,
Julia

> Dear colleagues,
>
> we have recently experienced severe problems with Dicty transformations
> when selection with G418 was required. We have used different plasmids
> with G418 cassettes, different AX2s, varied DNA concentrations, and
> transformation methods. We also ordered G418 from different sources and
> tried many G418 concentrations. Minimal G418 concentrations required to
> kill the cells seemed normal, yet we do not get clones. In parallel
> experiments transformation and selection with hygromycin or blasticidin
> works just fine. If anybody has overcome similar problems I would be eager
> to know the answer.
>
> Thanks a lot,
> Thomas
>
>
> -------------------------------------------------------
> Prof. Dr. Thomas Winckler
> Friedrich-Schiller-Universität Jena
> Institut für Pharmazie
> Lehrstuhl für Pharmazeutische Biologie
> Semmelweisstraße 10
> 07743 Jena, Germany
>
>
>


Dr. Julia von Bülow
Pharmazeutisches Institut
Christian-Albrechts-Universität zu Kiel
Gutenbergstr. 76
24118 Kiel
Tel: 0431 880-1812
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