Thesis (Ph. D.)--University of Adelaide, Dept. Plant Science, 1997. / Errata pasted on front fly-leaf. Includes bibliographical references (leaves 139-173).

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Errata pasted on front fly-leaf. Bibliography: leaves 139-173. This thesis identifies DNA structures and genes involved in the process of homologous chromosome pairing in allohexaploid bread wheat (Triticum aestivum). In addition to studying late replicating DNA, a speculative model on the action of the pairing genes in allohexaploid wheat and the putative function of the AWWM5 gene is discussed.

A two-component transposable element system consisting of a stabilized Activator (Acst) and a chimeric Dissociation (Ds) element has been introduced into the genome of Brassica napus. This Acst/ Ds system incorporates the use of several highly effective screenable and selectable markers. One of these markers is the maize Lc gene, a transcriptional regulator of flavonoid biosynthetic genes. This substrate-independent screenable marker was tested for the first time in B. napus and I show that when overexpressed, there is augmented trichome production and a light-dependent, enhanced accumulation of anthocyanins in B. napus plants. The phenotypes are expressed under a wide range of conditions, are visually distinct, and are observed throughout plant development. When used as a visual marker for the Acst element, Lc B. napus plants were rapidly identified among F2 segregating populations. As part of my goal to develop a very efficient Acst/Ds system for use in B. napus, a conditional negative selectable marker, the E. coli codA gene, was also tested for the first time in B. napus. This was done because use of a substrate-dependent negative selectable marker can facilitate the rapid and reliable identification of stable Ds transposition events when used as a marker for the Acst T-DNA. The enzyme cytosine deaminase, encoded by the codA gene, catalyzes the deamination of the non-toxic compound 5-fluorocytosine (5-FC) to the highly toxic compound 5-fluorouracil. In codA transformed B. napus seedlings, expression of cytosine deaminase results in a severe suppression of growth and this phenotype is dependent on the presence of the 5-FC substrate. Wild-type seedlings, however, lack endogenous cytosine deaminase activity and appear unaffected by the presence of 5-FC in the growth media. These results indicate that codA has the potential to be used effectively in B. napus as a substrate-dependent negative selectable marker for the Acst T-DNA. To determine if Ac transposase cou

Previous studies have shown that transcriptional differences in the orf224-atp6 mitochondrial gene region are correlated with fertility restoration of the pol CMS trait by the dominant nuclear Rfp gene in Brassica napus. Recently, the recessive rfp allele, or a tightly linked gene, was found to act as a dominant gene, designated Mmt, in controlling the production of additional, smaller transcripts of two other mitochondrial loci. The results presented in this thesis reveal that Mmt-specific transcripts lack sequences found at the $5 sp prime$ end of the full-length transcripts of these loci and contain a common sequence, UUGUGG, which maps immediately downstream of their $5 sp prime$ termini. A similar sequence, UUGUUG, is found within orf224 downstream of the major Rfp-specific $5 sp prime$ transcript terminus; these hexanucleotide sequences may serve as recognition motifs in the generation of Mmt- and Rfp-specific transcripts. These results suggest that Rfp/Mmt is a novel nuclear locus affecting the expression of multiple mitochondrial gene regions, with different alleles or haplotypes affecting different mitochondrial genes.

Cytoplasmic male sterility (CMS) in plants is associated with mitochondrial dysfunction. We have proposed in this study that the mitochondrial-encoded chimeric peptide thought to be responsible for cytoplasmic male sterility in Polima system could function as a dominant male sterility inducer when expressed in the nucleus and targeted to the mitochondria. Transgenic plants expressing such mitochondrial targeting constructs exhibited reduction of pollen production that was characterized in fertile Westar (nap ) and restored fertile Westar (pol) plants by homeotic transformation of floral organs and in male-sterile Westar (pol) plants by a reduction in pollen production with shortening of the stamens. Genetic and molecular analysis has shown that the phenotypic changes were correlated with the effective genetic transmission of the inserted transgene through female gametes. Most significantly, we have found that differences in floral morphology induced by transgene expression between pol CMS and fertile Westar plants might be related to differences in transcriptional activity of the APETALA3 MADS box gene. We suggest that the alterations in floral morphology that accompany CMS in several plant species might be due to effects of mitochondria on transcriptional activity of floral organ identity genes.

DNA markers tightly-linked to nuclear fertility restorer genes for cytoplasmic male sterility (CMS) are valuable tools for breeders and researchers working with these genes. Two different targeting approaches were used to identify markers linked to the Rfp1 restorer gene for the pol CMS of canola (Brassica napus L.): nearly isogenic line (NIL) comparison and bulked segregant analysis. These methods were equally efficient in identifying markers linked to Rfp1; combining them allowed a targeting efficiency of 100% to be achieved. The efficiency of bulked segregant analysis was found to be limited by the inadvertent occurrence of shared homozygosity at specific chromosomal regions in the bulks, in contrast with the efficiency of NIL comparison which was limited by the occurrence of residual DNA from the donor cultivar at scattered sites around the genome of the NILs. Eleven DNA markers linked to the Rfp1 gene were identified, one of which perfectly co-segregates with Rfp1. The linkage group on which Rfp1 is localized contains 17 DNA markers. Two restorer genes of the pol CMS, Rfp1 and Rfp2, and a Rfn restorer gene of the nap CMS were found to be at least tightly linked to one another and may all reside at the same locus. A fourth restorer gene, the Rfo restorer for the ogu CMS, was, however, found to be unlinked to the other restorer genes. Different restorer genes for the nap CMS were found in the lines 'Westar-Rf and 'Karat'. A linkage map of the B. napus genome containing 146 markers organized into 23 linkage groups covering a total length of 850.2 cM was constructed from a BC$ sb1$ population. This map contains 63 loci previously localized on the B. napus genome through analysis of an F$ sb2$ population. Comparative analysis indicates that the total length of the BC$ sb1$-derived map is smaller than that of the F$ sb2$-derived map, which suggests that a reduction in recombination frequency is occurring in male gametes. The preferential use of two or three probe-