International Mammalian Genome Society

17th International Mouse Genome Conference

9-12 November 2003, Braunschweig, Germany


POSTER 48 - MOUSE BAC CGH ARRAY TO IDENTIFY GERM-LINE MUTATIONS INDUCED BY HEAVY ION BEAM (HIB) IRRADIATION

Nagase H
Roswell Park Cancer Institute

Co-Authors: 1) Kimura MT, 1) Conroy JM, 1) Nowak N, 1) Igarashi J, 2) Yoshiki A, 3) Kusakabe M
Institutions: 1) Roswell Park Cancer Institute, 2) RIKEN BioResource Center, 3) ANB Tsukuba Institute

We have presented the heavy ion beam (HIB) approach of mutagenesis in previous IMGC meetings. There are unique advantages for HIB mutagenesis. Firstly, it uses a relatively low-dose, localized irradiation targeted specifically to the testis. Secondly, the mutation rate, as measured by the increased frequency of functional mutations (e.g. using HPRT), is lower than ENU mutagenesis. The frequency with which mutant phenotypes are produced is, however, similar. Thirdly, the mutations that are induced are mainly small deletions, which result from clustered double-strand DNA damage (Goodhead, 1994; Rydberg, 1996) . This is an important aspect of HIB, since it is subsequently relatively easy to detect deletions using molecular genome scanning approaches.

We collaborated with the Microarray & Genome Resource Core at RPCI and have recently developed mouse high resolution BAC CGH array. Ligation-mediated PCR was employed to generate representations of currently ~2,500 mouse BACs for printing with high reproducibility and provided essentially identical ratios as those generated using plasmid DNA from the same BACs. Details concerning DNA labeling and hybridization were reported previously (Snijders et al., 2001). The hybridized slides are scanned using an Affymetrix 428 scanner for both Cy3 and Cy5 channels. The output of the image analysis is processed by an in-house Perl program. The mapping data for each BAC is found by querying the mouse genome sequence at http://genome.ucsc.edu and the information is added to the resulting ratios and standard errors. We will discuss usefulness of this BAC array to detect germ-line deletions in HIB induced mutant mice.


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