Agrobacterium problems

at bio.tamu.edu at bio.tamu.edu
Thu Oct 12 18:56:42 EST 1995


In article <45jak8$8hm at highway.LeidenUniv.nl>, bundock at rulsfb.leidenuniv.nl
(Paul Bundock) wrote:

> In article 
> <Pine.OSF.3.90.951011100618.5908A-100000 at central.murdoch.edu.au>, 
> wylie at CENTRAL.MURDOCH.EDU.AU says...
> >
> >Does anyone else working with Agrobacterium tumefaciens have the 
> problems 
> >that I do?
> >Firstly, it is very inconsistant in growth, sometimes growing 
> vigorously 
> >in liqiud medium, and at other times growing slowly.
> >Transforming it with large plasmids by electroporation or triparental 
> >mating mating has been easy with some plasmids and impossible with 
> others 
> >of comparable size. All conditions and cells identical.
> >I use the strain Agli and have not tried others. Is there a difference 
> in 
> >ease of use between strains? Where can I buy other strains?
> >I store a glycerol stock at -80 C and plate onto LB + glucose every 
> >couple of weeks, working from single colonies taken from that plate. 
> >Any suggestions most welcome. Thanks.
> >Steve Wylie
> >Murdoch University
> >Western Australia

I haven't had your transfomation problems but I have seen your growth
problems and tracked the problem down to fresh plates.  If the agro plate
had been stored in the fridge at all then the cultures grow much slower.  I
have no idea why agro is so sensitive to chilling (maybe it has some sort
of dormancy reflex).  If you use freshly streaked plates then the cultures
should grow consitantly.  

I haven't noticed any strain to strain difference between competancy of
agrobacterium but I'm sure there is probably some variation.

As for buying agro strains, you can buy competant LBA4404 from BRL but I
think to get the other strains you'll have to write to labs that are using
them or that have stocks.  Since you're an Ausie I'd try contacting TJ
Higgins I think he's in Canberra, but there are several other labs around
Australia doing Agrobacterium transformation work and most of those will
have stocks of several different strains.

cheers
Bart



More information about the Methods mailing list