Abstract
The laboratory rat is rapidly gaining momentum as a mammalian genetic model organism. Although traditional forward genetic approaches are well established, recent technological developments have enabled efficient gene targeting and mutant generation. Here we outline the current status, possibilities and application of these techniques in the rat.
Highlights
Background mutationsAllows for allelic seriesTransposon-tagged mutagenesisGene insertions detectable by reporter gene cassettesRelatively low mutation efficiencyIntegration site easy to identifyBiased genomic integration pattern TargetedAllows gene targeting by nonhomologous end joining (NHEJ) and theoretically allows homologous recombinationModular assembly of zinc-finger arrays is relatively unsuccessfulHigh efficiency in introducing double-strand breaks (DSBs)Commercial zinc-finger nucleases (ZFNs) are expensive
To knock out genes in a targeted fashion without the need for pluripotent embryonic stem (ES) cells, one can use genetically engineered zinc-finger nucleases (ZFNs) [19]. This approach is based on the observation that double-strand breaks (DSBs), which are potentially lethal to the cell when they remain unrepaired, increase either homologous recombination and gene targeting or repair by error-prone nonhomologous end joining (NHEJ) [20]
If a DSB is introduced in the coding region of a gene or at an intron-exon boundary, repair by NHEJ can result in out-of-frame mutations or aberrant splicing and in a knockout allele
Summary
Contribute to the extensive phenotypic differences (including those relevant to common human disease) between these strains [11]. The availability of genome sequences of commonly used strains provides a useful resource to investigate the potential function and importance of genomic elements and polymorphisms that could be associated with disease states. Both forward (phenotypedriven) and reverse (genotype-driven) genetics approaches are instrumental to investigate such links between mutations and disease (see Figure 1). Alternative methods have been developed that enable efficient generation of mutants in a wide range of species The application of these techniques to the rat has resulted in the generation and characterization of a growing list of rat knockout animals that model human disease (Table 1)
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