Hi all!
I've been working with Arabidopsis plants from the past 2 years, however this is the first time I've noticed something strange. My plates containing T2 seeds have a white fungal contamination and I can't really place the issue. Some of the seeds have started giving rise to tiny plants and I really want to save them. Is there any way to do so? I had done plating about 5 days back. Thank you!
Will that be residue Agrobacterium? Some Agrobacterium can be carried to next generation.... Do you have a picture?
It looks like a cotton like white fungus to me. it has spread through my entire plates
Has the contamination spread to the seeds as well? If it has, it would be very hard to disinfect. Otherwise you can try to disinfect by using sodium hypochlorite and 70% ethanol. However that may affect the plantlets as well.
How did you disinfect the seeds before plating?
Abhishesh Bajracharya Hi, I usually sterilize the seeds using distilled water, 70% ethanol and mercuric chloride. I usually have no contamination in my plates using the technique I follow, However this time, I am observing a huge growth of a white coloured fungus. I shall try to use sodium hypochlorite this time too.
Dear Bipasha,
I am experiencing several contaminations after having sown my seeds onto gl. May you share the protocol you mentioned, by following which you have almost never contamination? Thanks a lot.
Davide
Davide Savy Hello! I usually sterilize my Arabidopsis seeds using the following method
1) Rinse the seeds 5-6 times with distilled water thoroughly.
2) Rinse with 70% ethanol once
3)Rinse with distilled water 2-3 times
4)Rinse with 0.05%Mercuric chloride once
5)Finally, thoroughly wash the seeds with distilled water 5-7 times
I perform the above steps in a laminar flow. I also make half MS plates a week in advanced to screen out plates which have contamination potential.
Hope it helps!
Hi Bipasha
you may use 5-20% bleach for 5 minutes, followed by 3-5 times washing with distilled water, for decontaminating your seeds. The seeds already contaminated shouldn’t be used further. Since you are working with T2 populatio, you are likely to have enough seeds.
all the best with your experiments.
Vivek
Vivek Dogra Thanks sir! It's a really helpful protocol that you've suggested. I've been using bleach of late replacing mercuric chloride and it's worked much better.
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