Division of Functional Genome Analysis


					Functional Genome Analysis (B070) Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 580 D-69120 Heidelberg, Germany.

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Tumour Analyses

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FINISHED PROJECTS:
Identification and characterisation of disease genes
by Representational Difference Analysis

Complementary and parallel to differential hybridisation techniques, Representational Difference Analysis (RDA) was employed as an alternative method for the detection of differentially expressed genes. The technique was adapted from the original protocol of Hubank and Schatz (1994) so that analyses are possible even with small amounts of starting material. In various projects, this technique was applied for the isolation of genes related with and potentially causative to diseases. In collaborations with the companies Merck, Hoffman la Roche and Knoll (now Abbott), analyses were carried out for the identification of disease-related genes on a wide variety of tissues. Large number of cancer-related transcriptional differences were identified and some of the relevant genes and gene products analysed in more detail. Also, antibodies were generated on the basis of such studies and are being used in actual protein expression analyses.

RDA was performed on larynx carcinoma tissue versus normal larynx tissue. Total difference products were cloned and and individula clones were picked. Redundancy of the library was minimised by iterative hybridisations of clones back to the library. PCR-products were spotted on filters, with the clones representing overexpression in normal and cancer tissue, respectively, being spotted in different orientation. Upon hybridisation of the respective starting material, all but one signal were of the expected orientation.

Gress et al.	.(1997)	Genes Chrom. Cancer 19, 97-103.
Geng et al.	.(1998)	Biotechniques 25, 434-438.
Frohme et al.	.(2000)	Ann. N.Y. Acad. Sci. 910, 85-104.
Frohme et al.	.(2002)	Biochem. Biophys. Res. Commun. 293, 1377-1382.
Scheideler & Hoheisel	(2002)	Screening 5/02, 22-25.
Zubakov et al.	.(2003)..	FEBS Lett. 547, 51-57.

Malanchi et al.	.(2004)..	J. Virol. 78, 13769-13778.
Frohme & Hoheisel	.(2006)..	Cell Biology 3rd ed., Elsevier, 113-120.
Steinberg et al.	.(2006)..	J. Periodontal. Res. 41, 426-446.
Schütz et al.	.(2006)..	J. Mol. Biol. 358, 997-1009.
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