Forward genetic screens for meiotic and mitotic recombination-defective mutants in mice.

Laura Reinholdt, Terry Ashley, John Schimenti, Naoko Shima

Research output: Contribution to journalArticlepeer-review

36 Scopus citations

Abstract

The goal of understanding the function of all mammalian genes is best accomplished through mutational analyses. Although the sequence of the mouse genome is now available and many genes have been identified, it is not possible to ascribe functions accurately to these genes in silico. Gene targeting using embryonic stem cells is ideal for analysis of individual genes selected on the basis of sequence features, but it is impractical for identifying novel genes involved in particular biological processes. Phenotype-based random mutagenesis of the genome is well suited for this goal. In the mouse, N-ethyl-N-nitrosourea (ENU) induces point mutations at a high frequency in the mouse germline. In this chapter, we describe methods for detecting and characterizing recombination mutations in mice produced by ENU mutagenesis. Potential meiotic recombination mutants are identified in a hierarchical fashion, by performing a screen for infertility, then gonad histology to determine whether meiotic arrest occurs, and finally by immunohistochemical analysis of meiotic chromosome with a battery of antibody markers. Screening for mutations potentially required for recombinational repair of DNA damage in somatic cells is performed using a flow cytometry-based micronucleus assay. Both strategies have proved effective in identifying desired classes of mutations.

Original languageEnglish (US)
Pages (from-to)87-107
Number of pages21
JournalMethods in molecular biology (Clifton, N.J.)
Volume262
StatePublished - 2004

Fingerprint Dive into the research topics of 'Forward genetic screens for meiotic and mitotic recombination-defective mutants in mice.'. Together they form a unique fingerprint.

Cite this