Hello everybody!
I wonder why T-DNA delivered by Agrobacterium in techniques like agroinfiltration is not integrated into the host genome, while other techniques like dipping are used for stable transformation. Is there any difference in the Agrobacterium strains or in the design of the vector?
Thanks in advance!
Hi Carlos,
As far as I am aware there are no specific strains that differentiate for these purposes. I have used Agrobacterium GV3101 (pSoup) successfully for floral dipping of Arabidopsis as well as transient transformations in both Arabidopsis and Nicotiana benthamiana
@Carlos:
As explained previously, there should be no Agro strain effect between the two approaches. My guess the only difference is probably associated with whether there is DNA replication in the Agro infected cells/tissues or not.
For example, in leaf Agro-infiltration - the agro-infiltrated leaf tissues did not undergo cell replication. Therefore, there is no DNA replication in the Agro-infected cells. Eventhough the introduced T strand actually reached the nucleoplasm, no T-DNA integration to the host chromosome occurs. The T-DNA integration probably occurs through non-homologous recombination, which requires the host DNA replication. In Agro-infiltration, the cells in leaf tissues are not dividing and neither DNA replication; therefore, there will be no integration of T-strand into host chromosome.
Moreover, since expression of the introduced transgene is independent of integration in the host chromosome, it is possible to do transient assay using Agro-infiltration.
On the other hand, in floral dip Agrobacterium mediated transformation, the target pollen mother cells are in the process of cell division to develop pollens (i.e. there is DNA replication). Therefore, when there are T-strand around in the nucleoplasm, T-strand integration into host chromosome through non-homologous recombination should be possible.
Good luck...
Hello Carlos!
While talking about plant, the compatibility of host and Agrobacterium strain can play an important role in transformation efficiency. Plant species have variable susceptibility to different Agrobacterium strains for both transient and constitutive T-DNA expression processes. However you can get enough information regarding comparison of both approaches from this review: (PDF) Agrobacterium-Mediated Transient Expression as an Approach to Production of Recombinant Proteins in Plants. Available from: https://www.researchgate.net/publication/23656743_Agrobacterium-Mediated_Transient_Expression_as_an_Approach_to_Production_of_Recombinant_Proteins_in_Plants [accessed May 09 2021].
The strains like LBA or EHA might not have exact roles as has been claimed.
There were indeed various factors that influence Agrobacterium t-DNA integration in host plants of which Agrobactrium strains, host plant receptivity, co-cultivation media, Agro concentration, temperature etc are quite important. For example, for some plant species, the EHA series may be better than LBA4404 while in some other species the reverse may be true. It is important how effectively you are making your Agrobacterium more virulent by manipulation. For more details, you can read papers available freely in researchgate.1. Agrobacterium-mediated transformation of coffee: An assessment of factors affecting gene transfer efficiency and 2. Two critical factors: Agrobacterium strain and Antibiotics selection regime improve the production of transgenic coffee plants.
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