Large-scale mapping of mutations affecting zebrafish development

Abstract

Background: Large-scale mutagenesis screens in the zebrafish employing the mutagen ENU have isolated several hundred mutant loci that represent putative developmental control genes. In order to realize the potential of such screens, systematic genetic mapping of the mutations is necessary. Here we report on a large-scale effort to map the mutations generated in mutagenesis screening at the Max Planck Institute for Developmental Biology by genome scanning with microsatellite markers.

Results: We have selected a set of microsatellite markers and developed methods and scoring criteria suitable for efficient, high-throughput genome scanning. We have used these methods to successfully obtain a rough map position for 319 mutant loci from the Tübingen I mutagenesis screen and subsequent screening of the mutant collection. For 277 of these the corresponding gene is not yet identified. Mapping was successful for 80 % of the tested loci. By comparing 21 mutation and gene positions of cloned mutations we have validated the correctness of our linkage group assignments and estimated the standard error of our map positions to be approximately 6 cM.

Conclusion: By obtaining rough map positions for over 300 zebrafish loci with developmental phenotypes, we have generated a dataset that will be useful not only for cloning of the affected genes, but also to suggest allelism of mutations with similar phenotypes that will be identified in future screens. Furthermore this work validates the usefulness of our methodology for rapid, systematic and inexpensive microsatellite mapping of zebrafish mutations.

Figure 1

Distribution of mapped mutations among the zebrafish chromosomes. Light blue, ENU mutations mapped in the present work. Purple, insertional mutations from the laboratory of N. Hopkins ([11][12] and unpublished data, available from ZFIN [10]), shown for comparison. Numbers for insertional mutations were obtained by searching ZFIN for mutations with a "hi" designation assigned to each linkage group and eliminating multiple hits of the same gene as well as mutations with ambiguous chromosomal assignments. Yellow, Ensembl genes predictions for each chromosome (× 100) (Ensembl release Zv6, available from Ensembl [13]) The number of mapped mutations or genes is indicated on the vertical axis, the linkage group (LG) number on the horizontal axis.