Rice Genetics IV - IRRI books - International Rice Research Institute

cesses. To study the interactions among genes, mutants in a particular pathway can becombined and the genetic hierarchy studied. These mutants can be precisely mappedand lead to gene isolation by map-based cloning procedures.Insertional mutagenesis, with transposable elements or Agrobacterium-mediatedT-DNA insertions, can generate mutants and also lead directly to gene identification.The insertion often causes a knockout mutation by blocking the expression of thegene and might display a mutant phenotype. The mutant gene tagged by the insertioncan be isolated by recovering DNA flanking the insert and subsequently lead to theisolation of the wild-type gene. For forward genetics, T-DNA insertional mutagenesisis only practical on a large scale for Arabidopsis (avoiding somaclonal variation) andhas resulted in a large number of tagged genes, although its use for reverse genetics inmodel plants such as rice has a future.The ability of the well-characterized maize transposon systems Ac-Ds and En-I(Spm) to transpose in heterologous hosts (Baker et al 1986, Pereira and Saedler 1989)offers new possibilities for transposon tagging, which is an efficient tool for identifyinggenes. For effective tagging strategies, two component systems comprising a mobiletransposon component (Ds or I/dSpm) and the corresponding stable transposase (Acor En/Spm) source have been developed (reviewed in Pereira 1998). The mobiletransposon components are often inserted in assayable/selectable marker genes tomonitor their excision phenotype (Baker et al 1987) and can also contain other convenientmarker/selectable genes to screen for their presence. To control transposition,the transposase is often put under the control of heterologous promoters (Swinburneet al 1992) or segregated out in progeny to yield stable transposon inserts. Astransposons move preferentially to closely linked sites (Jones et al 1990) and taggenes efficiently near their original position, efforts are made to map transposons inheterologous systems in order to generate jumping pads all over the genome for efficienttargeted mutagenesis.Ac-Ds transposons were first introduced by electroporation into rice (Izawa et al1991, Murai et al 1991, Shimamoto et al 1993) and shown to transpose. It was noted,however, that the two-component Ac-Ds system often gets inactivated after the firstgeneration (Izawa et al 1997), justifying a systematic investigation of Ac-Ds biologyin rice. Recently, the behavior of Ac has been followed through three successive generationsand it has revealed characteristics suitable for functional genomics strategies(Enoki et al 1999). Another knockout mutagenesis tool in rice, an endogenousretrotransposon, Tos17, has been developed for reverse genetics screens (Hirochika1997).The use of knockout mutations is limited, however, as the majority of genes displayno obvious phenotype (Burns et al 1994), probably because of functional redundancy,in which one or more other homologous loci can substitute for the same function.Even Arabidopsis, with its simple genome, contains large duplicated segmentsand redundant genes (Lin et al 1999, Mayer et al 1999). Therefore, the sequentialdisruption of redundant genes in an individual genotype might ultimately reveal amutant phenotype and uncover the gene function.Transposons and functional genomics in rice 265