Thesis (Ph. D.)--University of Hawaii at Manoa, 1994. / Includes bibliographical references (leaves [179]-196). / Microfiche. / xv, 196 leaves, bound ill. 29 cm

Typescript. / Thesis (Ph. D.)--University of Hawaii, 1969. / Bibliography: leaves [131]-135. / ix, 135 l graphs, tables

Stripe rust (Puccinia striiformis f. sp. hordei) is a serious disease of barley that can cause up to 70% yield loss in susceptible varieties. The fungus is moving northward, threatening major barley production areas in the US, where most cultivars are susceptible. Fungicides are available for control of stripe rust, but economic and environmental considerations favor genetic resistance. Two stripe rust resistance quantitative trait loci (QTLs) located in chromosomes 4 and 7 have previously been reported. One hundred and ten doubled haploid progeny from a stripe rust susceptible x resistant cross were derived using the Hordeum bulbosum technique and phenotyped for agronomic and malting quality traits in order to assess the importance of linkage drag associated with the mapped stripe rust resistance QTLs. Data on 33 markers were combined with phenotypic data for QTL analysis. A molecular marker-assisted backcross program was implemented to initiate the transfer of the stripe rust resistance loci into susceptible US germplasm. No negative QTLs for agronomic or malting quality traits were detected within or adjacent to the intervals that were targeted for marker-assisted selection. A minor leaf rust resistance QTL, however, was found adjacent to the stripe rust locus on chromosome 7. Linkage drag in this region could operate in favor of the breeder. Epistatic interaction between the two stripe rust resistance QTLs confirms the necessity of introgressing both chromosome intervals. / Graduation date: 1994

Black point of barley grain refers to a discolouration of the embryo end of the grain. Historically black point has been proposed to be due to fungal colonisation of the grain. However, Koch’s postulates have yet to be satisfied. The discolouration occurs during grain fill in response to high humidity or rainfall during the grain filling period. In wheat, which is also affected by black point, the discolouration has been proposed to be due to the oxidation of phenolic acids within the grain to form discoloured end products. Within this study, two approaches were investigated in order to understand the proteins and genes associated with this disorder. Firstly, a proteomics approach enabled the identification of individual proteins associated with black point. Two-dimensional gel electrophoresis was used to compare the proteome of the husk and whole grain tissue of mature black pointed and healthy grain. Very little watersoluble protein was extracted from the husk tissue. However, a significantly larger amount of protein was extracted using a salt extraction buffer, indicating the husk proteins were mostly cell wall bound. Due to the effect of residual salt and low protein concentrations these proteins were not conducive to analysis using two-dimensional gel electrophoresis. Further experiments using acid hydrolysis of the husk tissue and subsequent amino acid analysis revealed that the proteins were bound to the husk cell walls via covalent bonds. In contrast, large quantities of protein were obtained from the whole grain samples. This allowed statistically significant comparisons to be made between gels from healthy and black pointed grains. Two proteins were identified as being more abundant in black pointed grains. Mass spectrometry identified these as isoforms of barley grain peroxidase 1 (BP1). In addition, three proteins were identified as being more abundant in healthy grain. Mass spectrometry revealed these to be isoforms of the same protein with sequence similarity to a partial EST sequence from barley. Using 3' RACE the entire coding sequence of the gene was isolated which revealed that it encoded a novel putative late embryogenesis abundant (LEA) protein. Northern blot analysis was performed for BP1 and LEA and showed that gene expression differences could not account for the differences seen in protein quantities. Western blot analysis revealed that the LEA protein was biotinylated in vivo which is consistent with similar LEA proteins from other plant species. To further understand the role of these proteins in black point, antibodies were raised against the two proteins. Subsequent immunolocalisation studies indicated BP1 was present throughout all tissues of the grain whilst LEA was most abundant in the embryo and aleurone tissue. The second major area of investigation within this thesis was to further delineate the previously identified quantitative trait loci (QTL) associated with black point in barley. Previous studies have reported QTL for black point and kernel discolouration in both barley and wheat. Comparison of the published QTL revealed a locus on the short arm of chromosome 2H to be of particular interest. To identify genes underlying this QTL the genomes of barley, wheat and rice were compared. An in silico approach showed that the QTL shared macro-synteny with rice chromosomes 4 and 7. From the rice genome sequence, barley ESTs with sequence similarity were selected. In total, 20 ESTs were selected based on two main criteria: their putative role in black point and also being evenly spread across the region of the QTL length. These QTL were mapped within the Alexis x Sloop double haploid population. This approach revealed that there was some conservation of synteny but also identified clear boundaries where synteny between barley and rice had been lost since divergence. Significantly, the additional markers mapped to this region have enabled the initial black point QTL to be reduced from approximately 30cM to 20cM. In conclusion, this study has added significant knowledge our understanding of the genetics of black point in barley through the use of two approaches. The proteomics approach has aided in understanding the biochemical processes occurring within the grain in response to black point. The comparative genetics approach has aided in understanding the genetic control of an important region of the genome influencing black point susceptibility. Combined, these findings will direct future research endeavours aimed at producing black point resistant barley cultivars. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1323053 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2008

Abdominal Aortic Aneurysms (AAA) are amongst the top ten most common cause of death in those over 55 years of age. The disease is usually asymptomatic, often being diagnosed incidentally. Once diagnosed, elective repair of an AAA results in excellent long-term survival with a 3-5% operative mortality. However, up to one half of patients present with ruptured aneurysms, a complication that carries an 80% mortality in the community, and of those reaching hospital, a 50% mortality. Clearly early diagnosis and treatment results in improved survival. Screening for AAA, with ultrasound, would detect aneurysms early, prior to rupture. However, debate continues over the cost effectiveness of population based screening programmes. The identification of a sub-population at a higher risk of developing AAA would increase the yield of a screening prograrmne. A number of populations have been examined, none of which have received international acceptance. About 20% of patients with an AAA have a family history of an aneurysm. The disease is also considered to be a disease of Caucasians, both facts suggesting a strong genetic component to the disease. Perhaps a genetically identified sub-population at a high risk of developing an AAA would prove to be an ideal population for screening. This thesis examines the incidence of aneurysms and the family histories of patients with AAA in the Otago region of New Zealand. Almost twenty percent of the population has a family history of AAA. DNA was collected from each of these patients for genetic analysis. The population was divided into familial AAA and non-familial AAA for the purpose of genetic analysis and compared to a control population. AAA is believed to be a disease of Caucasians; a non-Caucasian population with a low incidence of AAA may prove to be a good control population for genetic studies. A literature review demonstrated a higher incidence of AAA in Caucasians than other ethnic groups and within Caucasians a higher incidence in patients of Northern European origin. The incidence was low in Asian communities, even in studies involving of migrant Asian populations. The New Zealand Maori are believed to have originated from South East Asia, therefore could be expected to have a low incidence of AAA and would make an ideal control population for genetic studies. A pilot study was undertaken to examine the incidence of AAA in the New Zealand Maori. The age standardised incidence of AAA proved to be at least equal in Maori to non-Maori, with a more aggressive form of the disease in Maori, manifesting with a younger age at presentation and a higher incidence of ruptured aneurysms at diagnosis. It is well known that at the time of surgery, an AAA is at the end stage in its life. At this time, inflammation and matrix metalloproteinases (MMP) enzymes are prevalent within the aneurysm wall and have destroyed the wall of the aorta. One of the most important genetic pathways regulating these enzymes is the plasminogen activator inhibiter 1-Tissue plasminogen activator-plasmin pathway. Genetic analysis of this pathway demonstrated an association of the 4G5G polymorphism in the promoter of the PAl-1 gene with familial AAA. In this insertion:deletion polymorphism, the 5G variant binds an activator and repressor, resulting in reduced PAI-1 expression and ultimately increased MMP activation. This allele was associated with familial aneurysms, 47% versus 62% non-familial AAA and 61% controls (p=0.024). A polymorphism within the tissue plasminogen activator gene was also examined and no association was found with AAA. Another way the MMPs expression could be increased is from mutations or polymorphisms in their own genetic structure. Stromelysin 3 is itself a MMP capable of destroying the aortic wall and it has a role in activating other MMPs. A 5A6A insertion:deletion polymorphism exists in the promoter of this gene. The 5A allele variant results in increased stromelysin expression and is associated with AAA 46% versus 33% in controls p=0. 0006. The actions of the MMPs are themselves inhibited by the tissue inhibitors of matrix metalloproteinases. The TIMP genes have been sequenced; two polymorphisms have been identified in the non-coding promoter area of the TIMP 1 gene. Further studies are necessary to examine the effect of these polymorphisms. Inflammation has been implicated in aneurysm progression. One of the roles of the inflammatory cells found in an aneurysm is to deliver the MMP�s to the AAA. The HLA system is integral in controlling this inflammation and was therefore examined. From this series of studies it is concluded that there is a genetic component to AAA. This thesis presents the first genetic polymorphism associated with familial AAA and explores the role of a genetic pathway in the formation of AAA.

Understanding the molecular basis of Mendelian disorders featuring joint dislocation can enhance the knowledge of genetic or cellular pathways required in joint development, and provide candidate genes for studying related complex disorders, such as developmental dysplasia of the hip. Two strategies were employed in this project to investigate Mendelian contributors to congenital joint dislocation. The first strategy was to investigate in-depth a gene known to be associated with joint dislocation. Missense mutations or small in-frame deletions in FLNB, encoding filamin B, have previously been associated with a spectrum of osteochondrodysplasias. Screening a larger cohort established FLNB as the sole underlying disease gene for atelosteogenesis type I and III and also boomerang dysplasia, which was previously thought clinically to be allelic to AOI. Mutations in FLNB cause a large proportion of Larsen syndrome cases with phenotypes reminiscent of the early case series reported. Atypical or "recessive" Larsen syndrome may therefore be due to a different underlying genetic aberration. The disease-associated amino acid substitutions or in-frame deletion/insertions cluster to two main regions of the filamin B protein: the calponin homology 2 domain of the actin-binding domain, and repeats 13-17 of the rod domain. To analyse the functions of these regions, yeast two-hybrid analyses were performed. No interactors were identified with the calponin homology 2 domain, which suggests the amino acid substitutions may disrupt actin binding or the regulation thereof. A candidate interactor, centromere protein J, was identified that binds to repeats 13-15, and could suggest a model for aberrant cell division seen in growth plates of bones of individuals with atelosteogenesis types I and III and boomerang dysplasia. The second strategy used in this project was to investigate the genetic cause of a novel syndrome featuring joint dislocation. A neurocutaneous phenotype segregated in a consanguineous New Zealand family, and through a genetic mapping strategy, a significantly linked locus was identified at 10q23 (Z = 3.63), in which segregation of a common ancestral haplotype fits the linkage hypothesis of homozygosity by descent. Candidate gene analysis and subsequent screening identified a missense mutation 2350C>T in ALDH18A1, which predicts the substitution H784Y in the encoded protein [Delta]�-pyrroline-5-carboxylate synthase (P5CS). The known function of P5CS in proline and ornithine biosynthesis was not affected by the presence of H784Y in an indirect assay, and therefore the hypothesis proposed was that a novel, unknown moonlighting function of P5CS is perturbed causing the phenotype segregating in the family. As an initial exploration of functions of P5CS in the cell, yeast two-hybrid analysis was undertaken. This project examined the contribution of two genes, FLNB and ALDH18A1, to Mendelian congenital joint dislocations. How the cellular functions of the encoded proteins in the cytoskeleton, metabolism, or signal transduction, are critical for joint development is ill understood. Future investigations aimed at identifying candidate genes that confer susceptibility to developmental dysplasia of the hip should consider candidate genes that encode proteins related in function to the products of the FLNB and ALDH18A1 genes.

The human immunodeficiency virus type 1 (HIV-1) uses a mechanism of genetic recoding known as programmed ribosomal frameshifting to translate the proteins encoded by the pol gene. The pol gene overlaps the preceding gag gene in the -1 reading frame relative to gag. It contains neither a start codon nor an internal ribosome entry site (IRES) to initiate translation of its proteins. Rather the host ribosomes are forced to pause due to tension placed on the mRNA when they encounter a specific secondary structural element in the mRNA. This tension is relieved by disruption of the contacts between the mRNA codons and tRNA anticodons at a �slippery� sequence within the ribosomal decoding centre. Re-pairing of the tRNAs occurs in the new -1 frame after movement of the mRNA backwards by one nucleotide, allowing the ribosome to translate the pol gene as a Gag-Pol polyprotein. A change in ratio of Gag to Gag-Pol proteins affects viral assembly, and most significantly dramatically reduces viral infectivity. The prevailing model for the mechanism of -1 frameshifting has focussed on a pre-translocational event, where slippage occurs when the slippery sequence is within the ribosomal A and P sites. This model precludes a contribution from the codon immediately downstream of the slippery sequence leading into the secondary structural element. I have termed this the �intercodon�. Often at frameshifting sites it is a termination codon, whereas in HIV-1 it is a glycine codon, GGG. When the intercodon within the frameshift element was changed from the wild-type GGG to a termination codon UGA, the efficiency of frameshifting decreased 3-4-fold in an in vivo assay in cultured human cells. This result mimicked previous data in the group within bacterial cells and cultured monkey COS-7 cells. Changing the first nucleotide of the intercodon to each of the three other bases altered frameshifting to varying degrees, but not following expected patterns for base stacking effects. Such a result would support a post-translocational model for -1 frameshifting. It suggested that the intercodon might be within the ribosomal A site before frameshifting, and that the slippery sequence was therefore within the P and E sites. This was investigated by modulating the expression of decoding factors for the intercodon - the release factor eRF1 and cognate suppressor tRNAs when it was either of the UGA or UAG termination codons, and tRNA[Gly] for the native GGG glycine codon. These were predicted to affect frameshifting only if slippage were occurring when the ribosomal elongation cycle was in the post-translocational state. Overexpression of tRNA[Gly] gave inconsistent effects on frameshifting in vivo, implying that its concentration may not be limiting within the cell. When eRF1 was overexpressed or depleted by RNAi, significant functional effects of decreased or increased stop codon readthrough respectively were documented. Expression of suppressor tRNAs increased readthrough markedly in a stop codon-specific manner. These altered levels of eRF1 expression were able to modulate the +1 frameshifting efficiency of the human antizyme gene. Overexpression of eRF1 caused significant reduction of frameshifting of the HIV-1 element with the UAG or UGA intercodon. Depletion of the protein by contrast had unexplained global effects on HIV-1 frameshifting. Suppressor tRNAs increased frameshifting efficiency at the UAG or UGA specifically in a cognate manner. These results strongly indicate that a post-translocational mechanism of frameshifting is used to translate the HIV-1 Gag-Pol protein. A new model (�almost� post-translocational) has been proposed with -1 frameshifting occurring for 1 in 10 or 20 ribosomal passages during the end stages of translocation, because of opposing forces generated by translocation and by resistance to unwinding of the secondary structural element. With translocation still incomplete the slippery sequence is partially within the E and P sites, and the intercodon partially within the A site. The nature of the intercodon influences frameshifting efficiency because of how effectively the particular decoding factor is able to bind to the partially translocated intercodon and maintain the normal reading frame.

Title page, contents and abstract only. The complete thesis in print form is available from the University of Adelaide Library. / Pre-harvest sprouting (PHS) is an important economic problem which affects a significant proportion of the Australian wheat crop through quality downgrading. Grain dormancy is the most effective means of overcoming germination in the wheat spikelet at harvest maturity. It has been a consistent observation over a long period of time that dormant red-grained wheat genotypes are almost more dormant than dormant white-grained genotypes. In white-grained wheat, there are two factors which contribute to dormancy, embryo sensitivity to abscisic acid (ABA) and an interacting and unknown seed coat factor. The proposed dormancy model is that complete dormancy can only be achieved with the coordinate expression of these two factors. This primary objective of this project was to determine the role of this putative seed coat factor in grain dormancy of white-grained wheat."--Abstract. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1259900 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2006

This thesis addresses the genetic basis of a rare autosomal recessive primary immunodeficiency disorder with the characteristic additional feature of venoocclusive disease of the liver (VODI). The interest in this condition was stimulated both by the potential to identify the genetic basis of a rare immunodeficiency and the opportunity to gain an insight into the biological basis of hepatic veno-occlusive disease, a poorly understood condition that is encountered most frequently in Australia as a consequence of bone marrow transplantation. The gene responsible for VODI was identified by homozygosity mapping and DNA sequence analysis of positional candidates and was shown to be the PML Nuclear Body expressed protein Sp110. This is the first time a PML Nuclear Body protein has been shown to be involved in immunodeficiency disorder. Subsequent immunofluorescence studies of affected patient cell lines showed absence of Sp110 in patient B cells. The role of SP110 alleles in the susceptibility of bone marrow transplant patients to hepatic veno-occlusive disease was investigated using a cohort of patients from the Fred Hutchinson Cancer Center, Seattle. A SNP association study identified initial evidence for an association, but the study lacked sufficient power after correction for multiple testing. Contemporaneously, Dr Igor Kramnik published a report that the murine homologue of Sp110, Ifi75 (also termed Ipr1) was deleted in mice that were supersusceptible to infection with Mycobacterium tuberculosis. A further SNP association study was therefore performed utilising a NSW cohort of Mantouxpositive South East Asian migrants, which detected evidence that alleles of SP110 may be associated with progression of M. tuberculosis infection. Again, the limited size of this cohort precluded definitive findings.

Bibliography : leaves 182-203. / 203 leaves : ill, map ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Describes the molecular analysis of the HLA class I and class II genes in the Australian Aboriginal population and also provides a comparison of serological and molecular tissue typing methods in view of genetic mutations at the HLA loci and the expression of serological HLA "blanks". / Thesis (Ph.D.)--University of Adelaide, Dept. of Medicine, 1996?