Dear All,
I am trying to generate over-expressing transgenic lines in Arabidopsis for my gene of interest by using Agrobacterium-mediated transformation. The protocol I followed for the floral dip was Steven J. Clough, Andrew F. Bent (Floral dip: a simplified method forAgrobacterium-mediated transformation of Arabidopsis thaliana).
However, no silique formation was observed in the transformed plants. I have repeated the experiment three times with around 30 plants but no silique formation was observed. However, the flowering was normal. What can be possible reasons for no silique formation?
Also, I have even tried to grow the deletion mutant lines ordered from NASC, even those mutant lines didn't even germinate.
It could be your Silwet, too much can damage the plants and they won't set seeds. Are you plants setting good seeds if they aren't dipped?
Thank you @Amy Klocko for your response. But I added Silwet as mentioned in the protocol and the same amount was added by another fellow lab mate. No such was observed in her case.
The non- transformed plants are setting good seeds.
Dear Pinky Dhatterwal,
If none of your NASC seeds germinate, I will try to order another mutant. However, if you find WT and He plants by PCR it could be that your mutant insertion line show embryo lethality.
Regarding to "no-seed" formation (and assuming that you are following the protocol properly), keep in mind that 35S promoter is poorly expressed in the anther tapetum so maybe your gene is essential for pollen formation. Thus, you can see flowers but not siliques because your embryos are aborted.
Hope you find it hepfull,
Check this publication from some collaborators_
Jesús Muñoz-Bertomeu, Borja Cascales-Miñana, Asunción Irles-Segura, Isabel Mateu, Adriano Nunes-Nesi, Alisdair R. Fernie, Juan Segura, Roc Ros, The Plastidial Glyceraldehyde-3-Phosphate Dehydrogenase Is Critical for Viable Pollen Development in Arabidopsis, Plant Physiology, Volume 152, Issue 4, April 2010, Pages 1830–1841, https://doi.org/10.1104/pp.109.150458
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