Is your problem that the plants are wilting, and/or drying out and
dropping leaves leading to death ?
If so, it is likely that you need to enclose the plants inside a
humid environment for a few hours (e.g. next day) after treatment.
Both the earlier vacuum treatment developed by Bechtold et al., and the
more efficient silwet transformation developed in Andrew's lab can lead to
a temporary breakdown of the plant's normal 'waterproofing'.
This can lead to extreme water loss through the leaves and any other
surfaces in contact with the atmosphere frequently followed by death in
dry or hot conditions.
The most convenient method of recovering from this is to drop the
plant pot (immediately after treatment) into a 'plastic sleeve' of the
kind available from florists or florist suppliers (also in supermarkets).
If you then allow the top of the sleeve to fold over, the plant is kept in
high humidity. A similar effect can be obtained with appropriate use of
small autoclave bags and masking/duck tape but is less elegant.
Please BE AWARE that you will need to open up the sleeve/bag after about a
day otherwise your plants will be prone to fungal infection and/or
sterility/developmental abnormalities. Direct sunlight may also cause
overheating in a closed bag so please be aware of this.
If this is not the problem, please describe the 'type' of death you see.
I hope this helps,
On 1 Aug 2000, zarir wrote:
>i am trying to transform arabidopsis plants with my promoter constructs
>in Agrobacterium tumefaciens GV3101 using the protocol on andrew bent's
>website. The first time i used the recommended 0.05% Silwet L-77 and my
>plants all died. The second time my plants died inspite of lowering the
>conc. of the silwet to 0.03%. Any suggestions to what i can change in
>the protocol from people with similar experience.
>N-304 Beadle Centre
>University of Nebraska-Lincoln
>Lincoln, NE 68588-0665
>Fax :- 402-472-3139
>Email:- zarir at purdue.edu>nematode at expert.cc.purdue.edu
Sean_May at nasc.nott.ac.uk (D.Phil.)
Director, Nottingham Arabidopsis Stock Centre (NASC)
School of Biosciences, Plant Science Division,
University of Nottingham, Biological Sciences Building,
University Park, Nottingham. NG7 2RD. UK
FON: +44 (0)115 9513237
FAX: +44 (0)115 9513297