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Several pathological conditions are caused by aberrant protein expression. To characterize the effect of abnormal protein expression, it is possible to employ transient overexpression models.
Moreover, a similar strategy can be employed to validate the pathogenetic role (or the biological function) of a protein. Indeed, overexpression experiments can be employed to rescue the phenotypes observed in a loss-of-function model. The injection of an mRNA encoding for the coding sequence (CDS) of interest can be employed to ubiquitously overexpress a protein during early zebrafish development (up to 3 days). Alternatively, to investigate the effect of protein overexpression at later stages or in restricted cell types, it is possible to inject a plasmid in which the CDS of interest is preceded by a ubiquitous or a tissue-specific promoter.
The expression of the exogenous CDS can be associated with one fluorescent reporter to facilitate the phenotypic analysis of the models generated.
Fast target validation prior to the generation of stable transgenic linesHigh throughput screening of multiple candidate genesNon-invasive, in vivo readouts to detect overexpression-induced phenotypes at the level of cells, tissues or organs
Fast target validation prior to the generation of stable transgenic lines
High throughput screening of multiple candidate genes
Non-invasive, in vivo readouts to detect overexpression-induced phenotypes at the level of cells, tissues or organs
To generate a model based on mRNA’s expression, zebrafish one-cell-stage embryos are injected with a solution containing the mRNA synthesized in vitro and encoding for the CDS of interest.
For plasmid overexpression, a donor plasmid containing the exogenous CDS of interest is injected into one-cell-stage embryos. During early stages of embryonic development, the plasmid can be imported in the nucleus and remain episomic. The exogenous CDS can be expressed by the promoter sequence included in the plasmid giving rise to F0 larvae transiently expressing the sequence of interest.
Results of the phenotypic analysis according to customers' interests and needs: