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Content archived on 2022-12-23

Expression studies using hybridisation-microchips with immobilised EST-oligonucleotides: identification of expressed genes in differentiated cells

Objective



Technology for the analysis by DNA by oligonucleotide hybridisation is being developed. A prototype for the sequencing by hybridisation on an oligomer-matrix (SHOM) has already been constructed consisting of microchips with gel-immobilised oligonucleotides and a specially designed fluorescence microscope, with a CCD-camera for detection, complete with image analysis software.

The hybridisation of fluorescence-labelled DNA to a SHOM microchip identifies oligonucleotide sequences contained in the DNA-fragment and the continuous sequence is determined from collation of these oligomer sequences. This technology will be used for the immobilisation of oligonucleotides that correspond to expressed sequence tags (ESTs), sequences which identify distinct genes. Such microchips will be applied for a quantitative determination of gene expression in different cell types and tissues. This will provide a way to produce atlases of expressed genes in differentiated cells and to establish the pattern of expressed genes involved in differentiation.

Such experiments might even allow regulative pathways to be defined following, for example, a sudden change in growth conditions. The small size of the microchips, the high concentration of gel-immobilised oligonucleotides, and the high level of fluorescence label of the probes will allow for the use of RNA isolated from small samples of tissues. At the beginning these microchips will be tested on known genes of Drosophila in comparison with results obtained from the analysis of cDNAs, then on more complex samples from, for example, Drosophila and C.elegans and finally on human, mouse and other higher organisms. Ultimately such a set-up could be used, for instance, as a system to test the biological effect/toxicity of various substances on a molecular basis.

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Coordinator

Deutsches Krebsforschungszentrum
EU contribution
No data
Address
Im Neuenheimer Feld 280
69120 Heidelberg
Germany

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Participants (2)

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