Dinoflagellate Gene Transformation
Did you catch that? If not check it out in slow motion:
Here is a still image of a GFP expressing Oxyhrris marina
What are Dinoflagellates?
What is Transformation?
Requirements for Gene Transformation:
First, a proper vector is required to carry the gene. Typically a plasmid is used. The plasmid carries the target gene, regulatory elements to drive the expression of the gene, and genes that will facilitate selection of the transformed cells (selectable marker) from non-transformed cells (needed because transformation efficiency is usually low), and a reporter gene that allows researchers to determine that transformation is successful.
Second, the vector needs to be delivered into the target cell. There is a range of methods, mainly electroporation, biolistic, and bacterium-mediated (infection, conjugation, or otherwise co-incubation).
Third, selection of the transformed cells require a chemical agent. One kind of selection agent would kill the non-transformed cells, but not the the transformed cells because the vector carries a resistance gene. Another kind of selection is by a source of essential nutrient (e.g. N) that can be utilized only by the transformed cells because they express an enzyme (e.g. nitrate reductase) metabolizing the source compound.
Funded by the Gordon and Betty Moore Foundation-Marine Microbiology Initiative (MMI), the over goal of this endeavor is to develop a transformation tool that is suited to transform dinoflagellates. This include two major objectives:
- Screen strains of dinoflagellates for one or more tractable candidates for stable transformation and genomic manipulation.
- Develop a protocol that can be applied to the dinoflagellate strain(s) identified.
This takes advantage of the fact that some dinoflagellates (especially heterotrophic and mixotrophic species) can take up bacteria.
Finding the right medium (low-salt) and suitable voltage is critical for success.
This requires that the species can grow on a solid (agar) medium.