Chapter 5 Genetic Analysis of Apomixis - cimmyt

mutations. Change of function ·mutations arethe result of amino acid substitutions or ofchanges in gene expression level. Loss offunction mutations abolish the gene productaltogether. As discussed earlier, apomixis maybe induced by a dominant allele, and this ismore likely to be the result of a change offunction mutation. There are numerous reportsof dominant mutations resulting from singleamino acid substitutions that have beenobtained by EMS or by in vitro mutagenesis(e.g., Hemerly et al. 1995; Kim et al. 1996;Wilkinson et al. 1997). For this reason, it couldbe important to use a mutagenic agent that caninduce subtle mutations. Ethylmethanesulfonate (EMS) has been shown to be the mostversatile mutagen, as it causes point mutationsat a high frequency, and whilst most of theseshould result in amino acid substitutions, theymay also create stop codons, and thus lead toloss of gene function (for a summary andreferences, see Feldmann et al. 1994). However,as already described, some useful mutantshave been obtained by radiation treatment,which frequently causes large deletions, andtherefore the value of mutagens that can causegene disruptions or deletions should not bedismissed.A widely used method of mutagenesis isinsertional inactivation, or gene tagging, withT-DNA or transposons (Feldmann 1991;Lindsey et al. 1993; Bouchez et al. 1993; Aartset al. 1995). As with deletions, this type ofmutagenesis most likely will create loss offunction mutations due to insertion into thecoding region of a gene, although it isconceivable that changes in gene expressionlevels could result from insertion into a genepromoter. However, a major advantage ofgenetagging is the ease with which the mutatedgene can subsequently be identified. Itsapplication to the study of apomixis inHieracium is examined by Bicknell (Chap. 8).We briefly summarize below the work byRamulu (1997) on transposon mutagenesis inArabidopsis and Petunia. For the purpose ofinducing apomixis in Arabidopsis, it may beimportant to choose a mutagen that is capableof producing change of function mutations.Arabidopsis is the best-characterized plantspecies, and map-based cloning of nontaggedmutant alleles is no longer a major obstacle.Some Early Work with MutantsInduction of Sexuality in ApomictsImproving cultivars of apomictic crop speciesby breeding is difficult, and therefore manyattempts have been made to increase thefrequency of sexual reproduction in apomictsby mutagenesis (Bashaw and Hoff 1962;Hanson and Juska 1962; Gustafsson and Gadd1965; Asker 1966). Complete reversion tosexuality was not observed in any of thesestudies. In the few cases where sexuality wasobserved, it was transient and plants revertedto apomixis in subsequent generations. Anumber of factors probably contributed to thislack of success. First, the induction of mutantsin polyploid species is inherently difficultbecause mutations can be masked byadditional copies of wild type genes. Second,in many experiments, the number of seedsmutagenized and screened (200 per treatmentin one case) was probably inadequate fordetecting potentially rare mutations. Whetherthe choice of mutagens, in most casesirradiaJion, also contributed to the lack ofsuccess depends on whether a change offunction rather than a loss of function mutationwould be required for a reversal to sexuality.We do not believe that the lack of success todate indicates that efforts to induce sexualityin apomictic species must necessarily fail.Researchers will need to work with a muchlarger number of mutants, to choose anapomict that has a low ploidy level, andperhaps to use a more subtle mutagenictreatment that induces both change and lossof function mutations.