Beyond the Identification of Transcribed Sequences: Functional and Expression Analysis

9th Annual Workshop, October 28-31, 1999

Co-sponsored by the U.S. Department of Energy


SIMONNEAU

Analysis of new transcribed sequences possibly involved in same functional pathways of neuronal development and identified by differential display

Michel Simonneau, Christophe Mas, Francine Bourgeois, and Fabien Guimiot

Neurogenetique/ INSERM E9935, Hopital Robert Debre, Paris, France

We identified new transcribed sequences using different differential display paradigms in order to select genes expressed in proliferating neuroblasts from mouse embryonic telencephalon. Differentially expressed genes were validated by radioactive relative RT-PCR. Their expression in proliferating telencephalon stem cells was tested by in situ hybridization on mouse embryo sections. From a set of 50 candidates, 58 % are unknown genes, whereas the other 42 % correspond to known genes. For these known genes, both their sequences and their pattern of expression suggest that they are involved in telencephalon stem cell proliferation.

We used similar approaches to isolate and validate differential expression of new genes either up or down regulated from in the developing brain of mouse mutant or knockout mouse embryos. Submicrogram of total RNA is sufficient to isolate and validate differential expression of up to 5 candidates.

As example, we found possible to identify new gene transcripts whose level varies in the brainstem as a function of a physiological variable (here, a ventilatory response), from Mash-1 +/- animals, suggesting that this approach is instrumental to identify new transcripts involved in a functional pathway.


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