Characterising genetic loci associated with loss of apomixis in Hieracium

Abstract

Most plant species strictly utilise sexual reproduction to generate genetically diverse seed to ensure adaptation of their descendents to the changing demands of their environment. Some species, however, have largely dispensed with sexual reproduction, opting instead to propagate clonally via apomixis, and maintain genotypes that are presumably already sufficiently adapted. Researchers of apomixis have long been attracted to the phenomenon as a biological curiosity, but more significant investigative attention is now being paid to it due to its ability to fix heterosis and therefore enable the economic production of high yielding hybrid varieties of the world's major crops.

Despite the strong motivation to integrate apomixis into seed-production systems, previous attempts to introgress the trait from wild apomictic relatives, or to induce it via mutagenesis, have yet to produce commercial apomictic varieties. It now appears likely that the successful transfer of apomixis into sexual crops will first require the elucidation of the molecular mechanisms, employed by native apomicts, that enable the avoidance of key components of sexual reproduction that otherwise serve to generate genetic diversity.

The Apomixis Programme at Crop & Food Research, Lincoln, aims to elucidate the genetics and molecular mechanisms of apomixis in Hieracium subgenus Pilosella. Two major deviations from sexual reproduction are required: the avoidance of meiosis, or apomeiosis, and the avoidance of fertilisation, or parthenogenesis. Segregating populations demonstrate independent segregation of apomeiosis and parthenogenesis. However, conventional mapping approaches towards determinants of apomixis in other species have often encountered significant difficulties posed by suppressed recombination at their loci. Alternative genetic resources of Hieracium were therefore generated using T-DNA and transposon mutagenesis, and deletion mutagenesis.

The present research focused on identifying and generating molecular maps of apomixes loci by screening deletion mutant panels of two genotypes, H. glaciale and H. caespitosum with secondary digest amplified fragment length polymorphism (SDAFLP). Identified loci were verified by their associations with apomixis in segregating populations, and SDAFLP markers were sequenced and converted into sequence characterised amplified regions (SCARs). The utility of the SCARs for the future isolation of BAC clones was determined by their presence or absence in key mutants. The identification and characterisation of three loci whose loss was associated with loss of -parthenogenesis in H. glaciale are described in Chapter 3. One locus transmitted to hybrid progeny as a determining locus and the other two transmitted as modifying loci. AT-DNA mutant of the H. glaciale background, which was included in the mutant panel, was found to carry a deletion at the determining locus. Findings that indicate that T-DNA insertions are not linked to the deletion are set out in Chapter 4, and somaclonal variation is suggested as an alternative cause of the deletion. Chapter 5 describes the use of deletion mutagenesis to identify two loci in H. caespitosum: one is associated with loss of apomeiosis (LOA) and the other with loss of parthenogenesis (LOP). Key mutants were screened with SDAFLP to obtain high densities of markers at LOA and LOP and markers that were predicted to be nearest the determinants were sequenced and converted into SCARs. One sequenced marker at LOP is likely to partially code for a regulatory gene. LOA and LOP segregated independently among hybrid progeny in strong association with apomeiosis and parthenogenesis respectively. Segregation distortion was characteristic of both loci, while recombination did not appear to be suppressed. Chapter 6 discusses how the findings of this research may be used to investigate the evolution of apomixis and to isolate its genetic determinants. It also discusses some challenges that might be encountered in the future during the engineering of apomixis in commercial crop species.