O aspartato é o precursor comum dos aminoácidos essenciais lisina, treonina, metionina e isoleucina. Devido à deficiência em lisina, a via metabólica do ácido aspártico tem sido estudada em cereais e os resultados obtidos indicaram a importância da aspartato quinase (AK), da homoserina desidrogenase (HSDH) e da dihidrodipicolinato sintase (DHDPS) como enzimas chave na regulação da síntese de lisina. Considerando-se a importância do milho e da quinoa como fontes de proteínas para humanos e animais, os objetivos deste trabalho foram: i) estudar as enzimas AK e HSDH nas sementes dos mutantes B77xB79o5; B37o7; W22o10, W22o11 e W22o13 e de seus respectivos tipos selvagens; ii) caracterizar a enzima DHDPS e analisar a composição de aminoácidos solúveis e incorporados nas proteínas das sementes dos mutantes Oh43o1, Oh43o2, Oh43fl1, Oh43fl2 e no respectivo tipo selvagem; iii) estudar nas sementes as enzimas AK, HSDH e DHDPS e alguns aspectos envolvidos na assimilação de nitrogênio e o perfil de aminoácidos de quinoa. Quando os mutantes B77xB79o5; B37o7; W22o10, W22o11 e W22o13 foram estudados, as análises demonstraram importantes variações nas atividades da AK e da HSDH entre os genótipos quando comparados com os respectivos tipos selvagens. A atividade da DHDPS foi fortemente inibida por lisina e não foram observadas variações significativas entre os genótipos Oh43o1, Oh43o2, Oh43fl1 e Oh43fl2, quando comparados com o genótipo Oh43+. A composição de aminoácidos revelou as maiores concentrações de lisina solúvel e incorporada em proteínas no mutante Oh43o2. Os estudos com quinoa demonstraram a presença de pelo menos duas isoenzimas da AK, uma sensível à inibição por lisina e a outra por treonina, duas isoenzimas da HSDH, uma resistente e a outra sensível à inibição por treonina e uma isoenzima da DHDPS sensível à inibição por lisina. As sementes de quinoa também apresentaram altas concentrações de lisina solúvel e incorporada em proteínas. / Aspartate is the common precursor of the essential amino acids lysine, threonine, methionine and isoleucine. Due to the deficiency in lysine, the aspartate metabolic pathway has been studied in cereal crops and the results obtained have indicated the importance of aspartate kinase (AK), homoserine dehydrogenase (HSDH) and dihydrodipicolinate synthase (DHDPS) as the key enzymes involved in the synthesis of lysine. Considering the importance of maize and quinoa as sources of protein for humans and animals, the objectives of this work were: i) to study the enzymes AK and HSDH in the seeds of the maize mutants B77xB79o5; B37o7; W22o10, W22o11 and W22o13 and their respective wild type; II) to characterize the enzyme DHDPS and to analyze the composition of soluble and amino acids incorporated into proteins in the seeds of the maize mutants Oh43o1, Oh43o2, Oh43fl1 and Oh43fl2 and the wild type OH43+; iii) to study the enzymes AK, HSDH, DHDPS in quinoa seeds and some aspects involved in nitrogen assimilation and amino acids profile. When the mutants B77xB79o5; B37o7: W22o10, W22o11 and W22o13 were studied, the analysis revealed important variations on AK and HSDH activities among the genotypes when compared to their respective wild type. The results indicated that DHDPS was strongly inhibited by lysine, and that there was not significant variation among the mutants Oh43o1, Oh43o2, Oh43fl1 and Oh43fl2 when compared to the wild type. The amino acids composition revealed high concentrations of lysine in the soluble form and incorporated into protein in the Oh43o2 mutant. The study of quinoa seeds allowed the identification of at least two AK isoenzymes, one sensitive to lysine inhibition and another sensitive to threonine, two HSDH isoenzymes, one resistant and another sensitive to threonine inhibition and one DHDPS isoenzyme sensitive to lysine inhibition. Quinoa also exhibited high concentration of soluble lysine and lysine incorporated into protein.

Title from container / Includes bibliographies and indexes / 4 v. : / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (D. Sc.)--University of Adelaide, Waite Agricultural Research Institute, 1987

Experiments were conducted to determine the outcrossing rate, the inheritance of markers and establish a basic linkage map in grasspea, <i> Lathyrus sativus </i>L. The outcrossing rate in a white-flowered line of grasspea ranged from 1.7 to 2.7% among eight combinations of gene frequency and location. The outcrossing rate in this study (2.2 ± 0.7%) suggests that individual lines of grasspea should be maintained in isolation to maintain their genetic integrity. Inheritance and linkage were determined for one morphological, 11 isozyme and 72 RAPD markers in five F₂ populations (all RAPD markers were in one F₂ population). The inheritance of flower colour was monogenic with colour dominant over white. The isozymes, ACO-1, ACO-2, AAT-1, AAT-2, EST-6, FDH, LAP-1, PGD-2, SKDH and TPI-1, were codominantly expressed with monogenic inheritance. The isozymes LAP-1 and PGD-2 segregated in a non-Mendelian ratios in the crosses PI 426891.1.3 x PI 283564c.3.2 and PI 426891.1 x PI 172930.4, respectively. The isozymeEST-3 was monogenically inherited and dominantly expressed. Most RAPD markers segregated in a 3:1 ratio. Marker UBC368_425/655 segregated in a co-dominant fashion. The RAPD markers UBC304₈₃₁, UBC304₉₆₄, UBC308₉₉₀, UBC322₁₄₃₂, UBC328₈₃₁, UBC332₁₁₁₈, UBC3321₁₅₈₁, UBC333₆₁₇, UBC349<sub>752<?sub>, UBC365₁₀₁₃ and UBC388₄₅₉ showed distorted segregation. In two F₂ populations, PI 283564c.3 x PI 426885.2 and PI 358601.5 x PI 173714.5, a linkage between AAT-2 and SKDH was reconfirmed. In the cross PI 426891.1.3 x PI 283564c.3.2, one morphological, three isozyme and 71 RAPD markers were mapped resulting in the delineation of 14 linkage groups including 69 markers (1 morphological, 3 isozyme and 65 RAPD markers). The total genome length covered by these 75 markers (69 linked and six unlinked) was about 864 cM. Considering cost, simplicity and abundance, RAPD analysis was more efficient than isozyme analysis in developing linkage map.

A basic understanding of the molecular events occurring during zygotic embryogenesis is required to fully understand how and why only a very small percentage of somatic embryos develop past the late embryogeny phase of embryogenesis. In this work, we have identified genes that have been demonstrated to be required for late embryonic development in the model plant system Arabidopsis thaliana. These genes were subsequently isolated and cloned from Loblolly pine embryos. These isolated clones were sequenced and analyzed to reveal significant homology to the known Arabidopsis ABA responsive genes ABI3, ABI4, and ABI5. Expression analyses of all three genes were completed, and compared to reported data of ABA accumulation, as well as, expression of other ABA responsive genes during the same stages of embryogenesis. Six putative root development genes were isolated and cloned from Loblolly pine embryos. These isolated clones were sequenced and analyzed to reveal significant homology to the known Arabidopsis root development genes WOODENLEG, SHORT ROOT, SCARECROW, HOBBIT, BODENLOS, and MONOPTEROS. Full-length cDNAs were isolated and cloned for WOODENLEG, SHORT ROOT, SCARECROW and BODENLOS. Expression analyses of all six genes were completed throughout mid to late embryogenesis in Loblolly pine.

Thesis (Ph. D.)--University of Chicago, Dept. of Molecular Genetics and Cell Biology, 2001. / Includes bibliographical references. Also available on the Internet.

Fenugreek (Trigonella foenum-graecum L.) is an annual legume mainly used as a spice crop in many parts of the world. "Tristar" is a new forage cultivar that requires - 120 days to produce mature seed in western Canada where only - 100 frost-free days are available. The goal for this study was to reduce maturity duration for the crop through a series of studies on the genetics and agronomic aspects of fenugreek. This two year study suggests that: 1)mutation breeding using Tristar seed as a base population could be successfull; 2)multi-location trials using world accessions exhibited genotype X environment interaction; 3)swathing of plants before freezing temperatures set in; 4)application of phosphate fertilizer increased seed yield and; 5)foliar sprays of chemicals can be used for production of high quality seed. In this study some short duration, high yielding and determine lines of fenugreek were produced improving the potential for use of fenugreek and the economics of beef production in western Canada. / xix, 184 leaves : ill. (some col.) ; 29 cm.

The genome of living organisms is constantly subjected to the environmental influences that result in different negative, negligible or positive impacts. The ability to maintain the genome integrity and simultaneously provide its flexibility is the main determinant for the evolutionary success of any species. One of the important aspects of genome maintenance is the precise regulation of the DNA repair machinery. Results reported here indicate the existence of a tight, age-dependent regulation of homologous recombination, one of the two main DNA double-strand break repair pathways. We show that recombination is influenced by conditions such as the change of temperature (cold or warm), day length, water availability (drought or overwatering stress) and salinity. These stresses not only influence the genome stability of stress-subjected generations but also change the recombination in subsequent generations. This indicates the possible involvement of homologous recombination in plant evolution and development of plant stress tolerance. / xiv, 121 leaves ; 29 cm.

In Arabidopsis thaliana (L.) Heyhn, the size of the pool of free proline increases up to 27-fold in response to osmotic stress. The magnitude of this accumulation is dependent upon the rate of imposition of the stress. Numerous reports have suggested a role for proline accumulation as a general adaptation to environmental stress. However, controversy surrounds the beneficial effect of proline accumulation in plants under adverse environmental conditions. Stress-induced proline accumulation in plants occurs mainly by de novo synthesis from glutamate. The final and only committed step of proline biosynthesis in plants is catalysed by Δ¹-pyrroline-5-carboxylate reductase (P5CR). The sequence of an incomplete 999 bp cDNA encoding P5CR from A. thaliana was determined. This enabled a preliminary molecular study of the structure and function of both the gene and the corresponding enzyme. The 999 bp cDNA insert in the clone Y AP057 was sequenced on the sense and antisense strands following subcloning of four sub-fragments in appropriate orientations. Comparison with known plant P5CR sequences revealed that Y AP057 does not encode the first 23 N-terminal amino acids of P5CR from Arabidopsis. However, it does encode the remaining 253 amino acid residues of Arabidopsis P5CR The cDNA Y AP057 is complete on the 3' end as indicated by the presence of a poly(A) tail. The nucleotide sequence determined shows complete homology to the corresponding exons of the genomic copy of a bona fide gene encoding P5CR in A. thaliana (Verbruggen et al, 1993). The only difference observed between the sequence of Y AP057 and that of a cDNA sequenced by these workers is that polyadenylation was initiated seven nucleotides earlier in Y AP057 than in the sequence of the published cDNA. Genomic Southern analysis suggests the presence of only a single copy of the gene encoding P5CR in Arabidopsis. Restriction mapping and sequencing the ends of another incomplete Arabidopsis P5CR cDNA clone FAFJ25 (664 bp) indicated that the regions sequenced were completely homologous to the corresponding portions of Y AP057. Analysis of codon usage in the Arabidopsis gene encoding P5CR revealed it to closely resemble the consensus pattern of codon usage in A. thaliana. This suggests that the gene is moderately. expressed. Expression of the gene encoding P5CR in Arabidopsis is not likely to be subject to translational control. Although P5CR from A. thaliana has a fairly high composition of hydrophobic amino acid residues, it does not possess any stretches of hydrophobic amino acids of sufficient length to act as membrane-spanning domains or to anchor the enzyme in a membrane. Neither does it contain an N- terminal leader sequence capable of directing it to either the plastid or mitochondrion. The enzyme therefore appears to be cytosolic. The nucleic acid and deduced amino acid sequences of Arabidopsis P5CR were compared with those from·eleven other organisms for which P5CR sequences are currently available. Except among the three different plants examined, P5CR sequences displayed less identity at the amino acid level than at the nucleotide level. The deduced amino acid sequence of Arabidopsis P5CR exhibits high similarity to the corresponding genes and amino acid sequences of P5CR from soybean and pea. Lower but significant similarity was observed to the amino acid sequences of P5CRs from human, Saccharomyces cerevisiae and the bacteria Escherichia coli, Pseudomonas aeruginosa, Thermus thermophilus, Mycobacterium leprae; Treponema pallidum and Methanobrevibacter smithii. Similarity was also observed to the translational product of a gene from Bacillus subtilis with high homology to the E. coli proC gene. However, construction of a phenogram indicating the relatedness of the various P5CR enzymes suggests that sequence analysis of this enzyme is not a good indicator of evolutionary relatedness of organisms from different biological kingdoms. Multiple alignment of the twelve known P5CR sequences indicated homology between the sequences across their entire lengths. Homology was particularly high in the C-terminal portions of the P5CRs studied. It is speculated that this region may be of importance in binding of the substrate Δ¹-pyrroline-S-carboxylate (P5C). Another region displaying high sequence conservation was found in the central portion of all P5CRs. All P5CRs studied, with the exception of PSCR from T. pallidum contained an N-terminal domain capable of binding a nicotinamide dinucleotide cofactor. Comparison of this region with consensus sequences for NADH and NADPH binding sites in proteins suggests that NADPH is the preferred reductant used by P5CRs from plants and human. In contrast, the N-terrninal domains of P5CRs from S. cerevisiae, M smithii, T. thermophilus and M leprae display greater similarity to a consensus NADH-binding site. The definite preference of plant P5CRs for NADPH in comparison with NADH suggests that P5CR may be involved in regulating the redox potential within plant cells and that this step in proline biosynthesis from glutamate may be of importance in overall metabolic regulation. Three amino acid residues are universally conserved in all P5CRs studied. All are found within blocks of high sequence similarity. These residues are likely to be of importance in the structure or catalytic mechanism of P5CR. A number of other residues are common to several of the enzymes examined. These may also be of importance in subsequent manipulation of Arabidopsis P5CR at the molecular level. Prediction of the putative secondary structures of A. thaliana, soybean, pea, human and E. coli indicated a high degree of similarity between the enzymes. This was particularly evident in the region of the putative P5C-binding domain. Considerable similarity exists in hydrophobicity profiles of P5CRs from these five organisms. Proline levels in reproductive organs of unstressed Arahidopsis plants were considerably higher than those in vegetative tissues. This suggests differential expression of enzymes involved in proline metabolism in these organs. In situ hybridisation studies indicated an increase in levels of mRNA transcripts encoding P5CR in stem tissues in response to water deprivation stress. Regulation of levels of mRNA transcript encoding P5CR in Arabidopsis therefore appears to be an osmotically sensitive process. Furthermore, this accumulation of transcript occurred in a tissue-specific manner. In particular, an increase in levels of transcript encoding P5CR was observed in the cortical parenchyma, phloem, vascular cambium and pith parenchyma in the vicinity of the protoxylem. The significance of these findings in contributing to a better understanding of the role of proline in adaptation to environmental stress is discussed. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1995.

Thesis (Ph. D.)--University of Missouri-Columbia, 1997. / Typescript. Vita. Includes bibliographical references (leaves 124-125). Also available on the Internet.

Thesis (Ph. D.)--University of Missouri-Columbia, 1997. / Typescript. Vita. Includes bibliographical references. Also available on the Internet.