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1
Rendell, Sarah. "Population genetic structure of Faidherbia albida (Del.) A. Chev. (Leguminosae, Mimosoideae)." Thesis, University of Oxford, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.299160.
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Hochschartner, Gerald. "Revealing the past : the potential of a novel small nucleolar RNA (snoRNA) marker system for studying plant evolution." Thesis, University of St Andrews, 2011. http://hdl.handle.net/10023/1695.
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Despite the existence of various molecular marker systems there are still limitations in distinguishing between closely related species based on molecular divergence, especially when hybridization events have occurred in the past. The characterisation of plant small nucleolar RNA (snoRNA) genes and their organisation into multigene clusters provides a potential nuclear marker system which could help in resolving the phylogenetic history of plants and might be applicable in DNA barcoding. Using closely and distantly related Senecio species, I investigated a combination of fragment length and sequence variation of snoRNA genes/snoRNA gene clusters to assess the utility of this marker system for barcoding and resolving species relationships. SnoRNA gene and gene cluster sequences identified in Arabidopsis thaliana were used to find homologues in other species and subsequently used for the design of universal primers. Most of the universal primer pairs designed were successful in amplifying snoRNA fragments in most Senecio species and fragment length variation between and within species could be detected. Furthermore, the combination of some fragment length datasets produced by different primer pairs enabled the separation of species and the detection of reticulate evolution indicating a high potential of snoRNA gene/gene cluster fragment length polymorphisms (SRFLPs) for phylogenetic reconstructions in Senecio and other plant genera. Most of the examined gene clusters showed a similar gene order in Senecio and Arabidopsis. However, the majority of these clusters appeared to exhibit more copies in Senecio, some of which were distinguishable by a combined sequencing/fragment profiling approach, and shown to be putative single copy regions with the potential to be used as co-dominant markers. However, a high number of paralogues and possible differences in copy number between species excludes these regions from being used in DNA barcoding. This is because specific primers would have to be developed for specific copies which would preclude development of a universal application for barcoding. None of the regions showed enough sequence variation to delimit distinctly closely related Senecio species and were therefore also considered to be unsuitable for DNA barcoding. Although most snoRNA genes and gene clusters might be inapplicable for DNA barcoding, they are likely to be valuable for phylogenetic studies of species groups, genera and families. On this scale, specific primers might act universally and the number of paralogous copies is likely to be equal across the species group of interest.
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Bitalo, Daphne Nyachaki. "Implementation of molecular markers for triticale cultivar identification and marker-assisted selection." Thesis, Stellenbosch : Stellenbosch University, 2012. http://hdl.handle.net/10019.1/71670.
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Thesis (MSc)--Stellenbosch University, 2012.Triticale is an amphidiploid that consists of wheat (A and B) and rye (R) genomes. This cereal is fast becoming important on a commercial basis and warrants further assessment for the better management and breeding of the hybrid. The assessment of the genetic diversity among the wheat and rye genomes within triticale can be obtained by using molecular markers developed in both donor genomes. Simple sequence repeats markers (SSRs) and amplified fragment length markers (AFLPs) have been previously used to assess the genetic diversity among triticale lines. SSRs are highly polymorphic markers that are abundant and which have been shown to be highly transferable between species in previous studies while AFLP markers are known to generate plenty of data as they cover so many loci. Thus, the aim of this study was to develop a marker system suitable to assess the genetic diversity and relationships of advanced breeding material (and cultivars) of the Stellenbosch University’s Plant Breeding Laboratory (SU-PBL). Therefore, both AFLP and SSR markers were initially analysed using eight triticale cultivars (with known pedigrees) to facilitate cultivar identification. Fourty-two AFLP primer combinations and 86 SSR markers were used to assess the genetic diversity among the Elite triticale cultivars. The AFLP primer combinations generated under average polymorphism information content (PIC) values. Furthermore, these markers generated neighbour-joining (NJ) and unweighted pair group method with arithmetic average (UPGMA) dendograms that displayed relationships that did not correspond with the available pedigree information. Therefore, this marker system was found not to be suitable. A set of 86 SSRs previously identified in both wheat and rye, was used to test the genetic diversity among the eight cultivars. The markers developed in wheat achieved 84% transferability while those developed in rye achieved 79.3% transferability. A subset of SSR markers was able to distinguish the cultivars, and correctly identify them by generating NJ and UPGMA dendograms that exhibited relationships that corroborated the available pedigree data. This panel of markers was therefore chosen as the most suitable for the assessment of the advanced breeding material. The panel of seven SSR markers was optimised for semi-automated analysis and was used to screen and detect the genetic diversity among 306 triticale entries in the F6, Senior and Elite phases of the SU-PBL triticale breeding programme. An average PIC value of 0.65 was detected and moderate genetic variation was observed. NJ and UPGMA dendograms generated showed no clear groupings. However, the panel of markers managed to accurately identify all cultivars within the breeding program. The marker panel developed in this study is being used to routinely distinguish among the advanced breeding material within the SU-PBL triticale breeding programme and as a tool in molecular-assisted backcross.
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Kisiel, John B. "Discovery and validation of aberrantly methylated DNA markers of pancreatic cancer." Thesis, College of Medicine - Mayo Clinic, 2015. http://pqdtopen.proquest.com/#viewpdf?dispub=1584252.
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Introduction: Pancreatic adenocarcinoma is anticipated to become the most fatal gastrointestinal neoplasm by 2017. Because patients who present with symptoms uniformly have advanced disease, 5-year survival is poor. Surveillance programs in the highest risk patients have produced disappointing results and use invasive tests that would not be safe or practical at the population level. Molecular markers, specifically aberrant DNA methylation, show promise in early studies. We aimed to: 1) measure the sensitivity and specificity of aberrant DNA methylation in stools as a minimally invasive way to detect pancreatic cancer; and, 2) identify novel methylation candidate markers highly sensitive and specific for pancreatic cancer.
Methods: To select candidate markers among those reported in the literature, DNA was extracted from unmatched tissue samples of pancreatic cancers and of normal colonic epithelia and assayed for aberrant methylation by methylation specific PCR. The most discriminant candidates and mutant KRAS were then assayed from matched archival stools of patients with pancreatic cancer in comparison to stools from healthy controls. Sensitivity and specificity for pancreatic cancer were determined from multivariate logistic regression models. To identify novel candidate markers, DNA was extracted from matched, archival tissues of pancreas cancer and two control groups (normal pancreas and normal colon) and sequenced using the reduced representation bisulfite technique. Among all mapped regions, those with the highest variance in methylation differences between cases and controls were filtered prior to analysis. Significant regions were then blindly assayed by methylation specific PCR in an independent, matched sample set, where sensitivity and specificity were measured using univariate logistic regression.
Results: In tissues, methylated BMP3, EYA4, UCHL1 and MDFI were highly discriminant for pancreatic cancer in comparison to normal colon samples. However, when assayed in stools, only BMP3 remained significant. In combination with mutant KRAS, the area under the receiver operating characteristics curve for BMP3 in detection of pancreatic cancer, was 0.85, indicating strong association. Results were not significantly influenced by tumor location or stage. Reduced representation bisulfite sequencing identified over 500 differentially methylated regions which met a priori significance thresholds. The top 25 novel candidates were validated in independent samples, showing both strong association with cases, compared to controls and high signal to noise ratio.
Conclusions: We report the first demonstration of feasibility for the detection of pancreatic cancer using assay of aberrantly methylated DNA markers from stool. This is a critical first step in the long-term goal of developing a minimally invasive screening tool to curb the mortality rate of this devastating disease. Because the discriminant candidate markers, methylated BMP3 and mutant KRAS, are not unique to pancreatic cancer, we developed a marker discovery strategy which yielded dozens of highly discriminant, validated, novel candidates, many of which have never before been reported in association with cancer. Further studies are indicated to measure the site-specificity of these markers for pancreatic cancer, compared to other gastrointestinal neoplasms and to study the clinical utility of these novel candidates in distant biologic media, such as blood, stool or pancreatic juice.
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Helgason, Thorunn. "Molecular markers in conservation genetics : chlorolast DNA variation in natural Scottish Pinus sylvestris L." Thesis, University of Edinburgh, 1993. http://hdl.handle.net/1842/14044.
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Scottish P. sylvestris populations have been extensively researched, using monoterpene and isozyme markers, but differentiation among populations is too low to identify origins or gene flow. Protocols were developed for the analysis of chloroplast DNA (cpDNA) Restriction Fragment Length Polymorphisms (RFLP) in P. sylvestris to determine whether cpDNA markers could be used for these purposes. Nine populations from throughout the range of pine in Scotland were sampled. Two individuals from each of these populations were surveyed using seven restriction enzymes and 13 probes from P. contorta, a total of 91 probe/enzyme combinations. The cpDNA genome of P. sylvestris was found to be about 119 kilobase pairs in length. 100% of the length of the genome was sampled, and 0.52% of the sequence length. No variation was found in any individual. These results were compared with a survey of P. sylvestris from China, Sweden and Turkey. There is no evidence that the cpDNA genotype of Scottish pine differs in any way from these varieties, suggesting that the cpDNA genome of P. sylvestris is homogeneous over a large part of the species' range. A survey of 191 individuals for 1 probe/enzyme combination revealed one variant individual, which appeared to be heteroplasmic for two cpDNA haplotypes. It was not possible to determine whether this was due to biparental inheritance of somatic mutation within that individual. The implications of these results for further research on the structure and inheritance of cpDNA in gymnosperms, the use of cpDNA as a genetic market in P. sylvestris, and the conservation of Scottish populations of this species are discussed. Finally, it is suggested that an increased collaboration between molecular biologists and ecologists is the best way to approach studies using sophisticated molecular techniques to measure genetic diversity in natural populations. In this way, the potential of this approach to improve the management of genetic resources can be fully realised.
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Pharmawati, Made. "DNA-based approaches for development of markers to assist Grevillea and Leucadendron breeding." University of Western Australia. School of Plant Biology, 2006. http://theses.library.uwa.edu.au/adt-WU2006.0110.
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[Truncated abstract] Grevillea and Leucadendron belong to Proteaceae and both have economic importance to the floriculture industry. Grevillea is a highly diverse genus endemic to Australia and very attractive for landscaping. Leucadendron is a South African Proteaceae but is cultivated in Australia and is well known as a cut flower. This thesis focuses on the application of DNA-based molecular markers to these genera. Several groupings within Grevillea were suggested by previous researchers based on morphological characteristics. In this thesis the monophyly of the groupings among 12 Grevillea species from New South Wales was tested using random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) analyses. To test the robustness of the data, UPGMA using Jaccard similarity, Neighbor Joining using total character difference and Wagner parsimony analyses were undertaken. The relationship trees generated supported monophyly of the groupings. Chloroplast DNA (cpDNA) was used to develop phylogenetic relationships among Leucadendron species. Inheritance and variation of cpDNA were evaluated using PCR-RFLP. The study demonstrated that cpDNA was inherited maternally and a phylogenetic tree of Leucadendron species using parsimony analysis was constructed. ... A fingerprinting study conducted using ISSR, produced a dendrogram showing the relationships among 30 cultivars. From the results, i a fingerprinting key was developed. Three examples of synonymous cultivar pairs were identified. In Leucadendron the male and female flowers develop on separate plants, and sex identification is only possible at time of flowering. ISSR, suppression subtractive hybridisation (SSH), and SSH combined with mirror orientation selection (MOS) were used in attempts of identifying sex-dependent DNA fragments at earlier stages of plant development. Neither of these techniques was able to identify sex-specific markers in Leucadendron. Nevertheless, the results did indicate that cpDNA copy number may differentiate male and female plants. Also, it was demonstrated that the genomes of male and female plants are quite homologous, which increases the difficulty in identifying sex-specific sequences. This thesis highlights the potential of DNA-based markers to determine species relationships in Grevillea and Leucadendron, as well as to identify Leucadendron cultivars. The information produced during the research for this thesis provides a basis for Grevillea and Leucadendron variety development and may be used to assist the design of interspecific crosses, to identify cultivars and the parents of hybrids. In addition, the results offer insights into the likelihood, problems and strategies of finding sex-specific markers for genes controlling sex in Leucadendron. ii
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Stenson, Andrew. "Use of molecular markers at different taxonomic levels : evolution of the northern lesser Antillean anole radiation." Thesis, Bangor University, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.327466.
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Drager, Robert Gray. "Molecular cloning of spinach chloroplast DNA isolated by alkaline lysis." PDXScholar, 1987. https://pdxscholar.library.pdx.edu/open_access_etds/3747.
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Chloroplast genomes of land plants show conservation of structure and gene arrangement. The spinach chloroplast genome is comprised of a covalently closed. circular DNA molecule of 150 kilobases and is typical of these plants. Approximately 20% of the proteins found in the spinach chloroplast are encoded by the chloroplast genome and translated on chloroplast ribosomes. The remainder are encoded on chromosomes in the nucleus, translated on cytoplasmic ribosomes and transported into the chloroplast.
Spinach chloroplast DNA was isolated from crude 2 chloroplast preparations by a new method. Chloroplasts were lysed with alkaline sodium dodecyl sulfate, contaminating macromolecules precipitated with acidified potassium acetate and plastid DNA was purified by phenol:chloroform extraction and ethanol:ammonium acetate precipitation. The yield was approximately 50 ug chloroplast DNA per 100 grams leaf material. The DNA consisted of 10% circular molecules and 90% linear molecules.
The chloroplast DNA was digested with restriction enzyme PstI and the fragments were cloned into the plasmid vector pUC9. Several recombinant plasmids were isolated and the chloroplast DNA inserts identified. The recombinant plasmid pRD105 containing the PstI #5 fragment was subjected to further investigation. The ClaI restriction sites of the PstI #5 fragment were mapped and the insert was subcloned into the plasmid vector pGEM4, which bears bacteriophage SP6 and T7 RNA polymerase promoter sequences.
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Vasemägi, Anti. "Evolutionary genetics of Atlantic salmon (Salmo salar L.) : molecular markers and applications /." Umeå : Dept. of Aquaculture, Swedish Univ. of Agricultural Sciences, 2004. http://epsilon.slu.se/s324.pdf.
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Fotinos, Tonya D. "Genetic Structure of the Florida Key Tree Cactus, Pilosocereus robinii, using Restriction Site associated DNA (RAD) markers." FIU Digital Commons, 2013. http://digitalcommons.fiu.edu/etd/914.
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Rare plant conservation efforts must utilize current genetic methods to ensure the evolutionary potential of populations is preserved. One such effort involves the Key Tree Cactus, Pilosocereus robinii, which is an endangered columnar cactus native to the Florida Keys. The populations have precipitously declined over the past decade because of habitat loss and increasing soil salinity from rising sea levels and storm surge. Next-generation DNA sequencing was used to assess the genetic structure of the populations. Twenty individuals representative of both wild and extirpated cacti were chosen for Restriction Site Associated DNA (RAD) analysis. Samples processed using the HindIII and NotIII restriction enzymes produced 82,382,440 high quality reads used for genetic mapping, from which 5,265 Single Nucleotide Polymorphisms (SNPs) were discovered. The analysis revealed that the Keys’ populations are closely related with little population differentiation. In addition, the populations display evidence of inbreeding and low genetic diversity.
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Arnold, Amanda Louise. "Development of molecular markers for studying population structure in marine fishes : 'coding versus non-coding DNA' : which is best?" Thesis, University of Aberdeen, 2000. http://digitool.abdn.ac.uk/R?func=search-advanced-go&find_code1=WSN&request1=AAIU485291.
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This thesis set out to assess whether variants identified within either the "coding" or the "non-coding" regions of the genome were more effective in resolving population structure in marine fishes. The assessment was carried out on three marine fish species - haddock (Melanogrammus aeglefinus), the lesser sandeel (Ammodytes marinus) and the Atlantic salmon (Salmo salar). The study was extended to compare variation within the "coding and "non-coding" regions of a section of the transferrin gene directly with nucleotide sequencing of the cDNA isolated from haddock and sandeels. Variants were identified both the "coding" and "non-coding" regions of this section of the transferrin gene selected and a population analysis was carried out to assess which source of variants was more effective for resolving population differences. Overall, the findings of this study were inconclusive. In some cases variation identified within coding regions was better than variation identified in non-coding regions of the transferrin gene for resolving population structure. However, in other cases the opposite was true, or both sources of variation were found to be uninformative with regards to genetically differentiating between populations. This inconclusiveness was partially attributable to the small number of variants that were identified in coding region for both haddock and Atlantic salmon and the fact that variation within only a single gene was studied. This study did, however, indicate that the methodology employed for screening individuals has clear advantages over previous methods used for population analysis, e.g. protein electrophoresis and mini/microsatellite analysis. All the variants present in the transferrin gene were detected using nucleotide sequencing. In contrast, using protein electrophoresis only those variants that cause an amino acid change and can be detected using histochemical stains are revealed. Difficulties were, however, experienced in accurately assigning allele sizes using mini- and microsatellites and led to binning of the data. This had the disadvantage that by increasing the accuracy of the data set, much of the resolution is likely to be lost.
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Campeol, Nadia. "Detection of markers in a low-density region of the barley (Hordeum vulgare L.) genome and their effects on the mapping of quantitative traits." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape10/PQDD_0002/MQ44137.pdf.
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Pena, Michelle Mendonça [UNESP]. "DNA Barcoding em Utricularia (Lentibulariaceae)." Universidade Estadual Paulista (UNESP), 2015. http://hdl.handle.net/11449/136705.
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000859555.pdf: 4094738 bytes, checksum: 1c3690c0cf5379b39472b8236e1cdb9c (MD5)A família Lentibulariaceae Rich. é considerada o maior grupo de plantas carnívoras dentre as angiospermas. Utricularia é o gênero de maior riqueza, com aproximadamente 250 espécies. Diversos estudos de identificação baseados em morfologia foram realizados para a família Lentibulariaceae, porém eles se mostram limitados para determinados grupos de espécies. Com base nisso a aplicação do DNA Barcoding pode ser uma importante alternativa. No presente estudo foram utilizadas sequências de DNA dos espaçadores intergênicos cloroplastidiais trnS-trnG e trnL-trnF e também do gene mitocondrial coxI com o objetivo de testá-las com a abordagem DNA Barcoding na diferenciação intraespecífica, interespecífica e entre as seções do gênero Utricularia. Com base nas matrizes de distâncias, a distância intraespecífica média foi de 0,004 para ambos os marcadores cloroplastidiais e de 0,006 para o gene coxI, a distância interespecífica média foi de 0,260 para trnS-trnG, 0,190 para trnL-trnF, 0,043 para coxI e a distância média entre as seções foi de 0,036, 0,029 e 0,025 para trnS-trnG, trnL-trnF e coxI, respectivamente. A análise baseada na árvore de Neighbor-Joining indicou que a maioria das espécies se agruparam em seções de acordo com o proposto para a filogenia do gênero, formando grupos monofiléticos. A eficácia de discriminação interespecífica foi 82% para trnS-trnG e 61% trnL-trnF, a discriminação intraespecífica foi de 36% para trnS-trnG e 23% para trnL-trnF. O gene mitocondrial coxI apresentou 24% de discriminação inter e intraespecífica, com resolução baixa de espécies na árvores de Neighbor-Joining. Esses resultados demonstram que as regiões cloroplastidiais apresentam informações satisfatórias para separação das espécies em clados que corroboram com a filogenia do grupo e que portanto trnS-trnG e trnL-trnF podem ser considerados bons barcodes para o...
The family Lentibulariaceae Rich. is considered the largest group of carnivorous plants among the angiosperms. Utricularia is the richest genus with approximately 250 species. Several studies based on morphological identification have been published for the family Lentibulariaceae, but they are limited regarding some groups of species. Hence, DNA Barcoding may be an important alternative. The present study used DNA sequences of chloroplast intergenic spacers trnS-trnG and trnL-trnF and also the mitochondrial gene coxI in order to test them with the DNA Barcoding approach to intraspecific, interspecific and between sections differentiation in the Utricularia genus. Based on the distance analyses, the average intraspecific distance was 0.004 for both chloroplast markers and 0.006 for the coxI gene, the average interspecific distance was 0.260 to trnS-trnG, 0.190 to trnL-trnF, 0.043 to coxI and the average distance between sections was 0.036, 0.029 and 0.025 to trnS-trnG, trnL-trnF and coxI, respectively. The analysis based on Neighbor-Joining tree indicated that most species were grouped into sections according to the proposed for the phylogeny of the genus, forming monophyletic groups. The efficacy of interspecies discrimination was 82% to trnS-trnG and 61% to trnL-trnF, intraspecific discrimination was 36% to trnS-trnG and 23% to trnL-trnF. The mitochondrial gene coxI showed 24% of inter and intraspecific discrimination, with low resolution of species on trees Neighbor-Joining. These results demonstrate that the chloroplast regions have satisfactory information to separate species in clades that corroborate the phylogeny of the group and therefore trnS-trnG and trnL-trnF can be considered good barcodes for the Utricularia genus
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Pena, Michelle Mendonça. "DNA "Barcoding" em Utricularia (Lentibulariaceae) /." Jaboticabal, 2015. http://hdl.handle.net/11449/136705.
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Orientador: Vitor Fernandes Oliveira de MirandaCoorientador: Alessandro de Mello Varani
Banca: Marcos Tulio de Oliveira
Banca: Yoannis Domínguez Rodríguez
Resumo: A família Lentibulariaceae Rich. é considerada o maior grupo de plantas carnívoras dentre as angiospermas. Utricularia é o gênero de maior riqueza, com aproximadamente 250 espécies. Diversos estudos de identificação baseados em morfologia foram realizados para a família Lentibulariaceae, porém eles se mostram limitados para determinados grupos de espécies. Com base nisso a aplicação do DNA "Barcoding" pode ser uma importante alternativa. No presente estudo foram utilizadas sequências de DNA dos espaçadores intergênicos cloroplastidiais trnS-trnG e trnL-trnF e também do gene mitocondrial coxI com o objetivo de testá-las com a abordagem DNA "Barcoding" na diferenciação intraespecífica, interespecífica e entre as seções do gênero Utricularia. Com base nas matrizes de distâncias, a distância intraespecífica média foi de 0,004 para ambos os marcadores cloroplastidiais e de 0,006 para o gene coxI, a distância interespecífica média foi de 0,260 para trnS-trnG, 0,190 para trnL-trnF, 0,043 para coxI e a distância média entre as seções foi de 0,036, 0,029 e 0,025 para trnS-trnG, trnL-trnF e coxI, respectivamente. A análise baseada na árvore de "Neighbor-Joining" indicou que a maioria das espécies se agruparam em seções de acordo com o proposto para a filogenia do gênero, formando grupos monofiléticos. A eficácia de discriminação interespecífica foi 82% para trnS-trnG e 61% trnL-trnF, a discriminação intraespecífica foi de 36% para trnS-trnG e 23% para trnL-trnF. O gene mitocondrial coxI apresentou 24% de discriminação inter e intraespecífica, com resolução baixa de espécies na árvores de "Neighbor-Joining". Esses resultados demonstram que as regiões cloroplastidiais apresentam informações satisfatórias para separação das espécies em clados que corroboram com a filogenia do grupo e que portanto trnS-trnG e trnL-trnF podem ser considerados bons "barcodes" para o...
Abstract: The family Lentibulariaceae Rich. is considered the largest group of carnivorous plants among the angiosperms. Utricularia is the richest genus with approximately 250 species. Several studies based on morphological identification have been published for the family Lentibulariaceae, but they are limited regarding some groups of species. Hence, DNA Barcoding may be an important alternative. The present study used DNA sequences of chloroplast intergenic spacers trnS-trnG and trnL-trnF and also the mitochondrial gene coxI in order to test them with the DNA Barcoding approach to intraspecific, interspecific and between sections differentiation in the Utricularia genus. Based on the distance analyses, the average intraspecific distance was 0.004 for both chloroplast markers and 0.006 for the coxI gene, the average interspecific distance was 0.260 to trnS-trnG, 0.190 to trnL-trnF, 0.043 to coxI and the average distance between sections was 0.036, 0.029 and 0.025 to trnS-trnG, trnL-trnF and coxI, respectively. The analysis based on Neighbor-Joining tree indicated that most species were grouped into sections according to the proposed for the phylogeny of the genus, forming monophyletic groups. The efficacy of interspecies discrimination was 82% to trnS-trnG and 61% to trnL-trnF, intraspecific discrimination was 36% to trnS-trnG and 23% to trnL-trnF. The mitochondrial gene coxI showed 24% of inter and intraspecific discrimination, with low resolution of species on trees Neighbor-Joining. These results demonstrate that the chloroplast regions have satisfactory information to separate species in clades that corroborate the phylogeny of the group and therefore trnS-trnG and trnL-trnF can be considered good barcodes for the Utricularia genus
Mestre
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Soleimani, Vahab D. "Genome dynamics in barley (Hordeum vulgare L) cultivars: Molecular diversity, evolution, and DNA fingerprinting." Thesis, University of Ottawa (Canada), 2005. http://hdl.handle.net/10393/29264.
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In the absence of whole genome sequencing, molecular markers are indispensable tools for the study of genome evolution, genetic diversity measurements, and genotype identification. We have used sequence-specific amplified polymorphism (S-SAP) markers that were derived from the BARE-1, an active retrotransposon of barley, to measure the contribution of this element to the evolution of barley genome among 103 cultivars that are commonly grown in Canada and the United States. The results were compared to the genome diversity measures obtained by the single nucleotide polymorphisms (SNP).
The barley populations were divided into groups based on various agronomic traits such as end use, i.e., feed versus malting, and seed morphology, i.e., naked versus covered kernel. Analysis of the genetic structure in the population using analysis of molecular variance (AMOVA) for both S-SAP and SNP attributed the largest co-variance component (90%) to the genetic diversity among cultivars within groups. Co-variance component between groups was about 6% which indicated that there was no justification for population differentiation along the set based upon agronomic traits. Genetic relationships among cultivars was assessed by cluster analysis with UPGMA and found to vary substantially between S-SAP and SNP datasets.
Quantitative analysis of BARE-1 retrotransposon with real-time PCR in a small group of cultivars showed significant differences in the copy number of the element among cultivars. Most of the BARE-1 elements were in the form of solo LTRs, indicating a high rate of homologous recombination between retrotransposon copies in the genome. Differences of up to 3000 BARE-1 copies per haploid genome were found among cultivars that have been developed and registered within the past three decades. Informative SNPs such as those with high polymorphic information content (PIC) values were used to generate identification keys to distinguish barley cultivars which were otherwise indistinguishable at the morphological and biochemical levels.
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Hu, Jiahuai. "Phytophthora nicotianae: Fungicide Sensitivity, Fitness, and Molecular Markers." Diss., Virginia Tech, 2007. http://hdl.handle.net/10919/26416.
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Mefenoxam has been a premier compound for Phytophthora disease control in the nursery industry for 30 years. The primary objectives of this research were to examine whether Phytophthora species have developed resistance to this compound and to investigate fungicide resistance management strategies. Phytophthora nicotianae, a destructive pathogen of numerous herbaceous and some woody ornamental plants, was used as a model system. P. cinnamomi, a major pathogen of a wide range of tree species and shrub plants, was also included for comparison.
Twenty-six isolates of P. nicotianae were highly resistant to mefenoxam with a mean EC50 value of 326.5 µg/ml while the remaining 70 were sensitive with an EC50 of <0.01 µg/ml (Label rate: 0.08µg/ml). All resistant isolates were recovered from herbaceous annuals and irrigation water in 3 Virginia nurseries. Resistant isolates were compared with sensitive ones using seedlings of Lupinus â Russell Hybridsâ in the absence of mefenoxam for relative competitive ability. Resistant isolates out-competed sensitive ones within 3 to 6 sporulation cycles. Resistant isolates exhibited greater infection rate and higher sporulation ability than sensitive ones.
No mefenoxam resistant isolates were identified in P. cinnamomi. All 65 isolates of P. cinnamomi were sensitive to mefenoxam with an EC50 of < 0.04 ï g/ml. Attempts to generate mutants with high resistance to mefenoxam through UV mutagenesis and mycelial adaptation were not successful. However, there were significant reductions in sensitivity to mefenoxam; those slightly resistant mutants carried fitness penalties, which may explain why P. cinnamomi remains sensitive to mefenoxam.
The effect of propamocarb hydrochloride on different growth stages of Phytophthora nicotianae was evaluated in search for an alternative fungicide. Propamocarb greatly inhibited sporangium production, zoospore motility, germination and infection. However, it has little inhibition of mycelial growth and infections. Propamocarb can be used as an alternative fungicide to mefenoxam where mefenoxam resistance has become problematic. However, it must be used preventively; i.e. before infections occur.
The genetic inheritance of mefenoxam resistance in P. nicotianae was studied using F1 progenies of a cross between resistant and sensitive isolates. The F1 progenies segregated for mefenoxam resistance in ratio of 1R:1S, indicating the mefenoxam resistance is controlled by a single dominant gene. One RAPD marker putatively linked to resistant locus in repulsion phase was obtained by bulked segregant analysis and was converted to the SCAR marker. This marker is capable of differentiating mefenoxam resistant populations from sensitive populations included in this study.Ph. D.
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Soto-Calderon, Ivan D. "Evolution of Nuclear Integrations of the Mitochondrial Genome in Great Apes and their Potential as Molecular Markers." ScholarWorks@UNO, 2012. http://scholarworks.uno.edu/td/1510.
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The mitochondrial control region (MCR) has played an important role as a population genetic marker in many taxa but sequencing of complete eukaryotic genomes has revealed that nuclear integrations of mitochondrial DNA (numts) are abundant and widespread across many taxa. If left undetected, numts can inflate mitochondrial diversity and mislead interpretation of phylogenetic relationships. Comparative analyses of complete genomes in humans, orangutans and chimpanzees, and preliminary studies in gorillas have revealed high numt prevalence in great apes, but rigorous comparative analyses across taxa have been lacking.
The present study aimed to systematically compare the evolutionary dynamics of MCR numts in great apes. Firstly, an inventory numts derived from the region containing the MCR subdomains was carried out by genomic BLAST searches. Secondly, presence/absence of each candidate numt was determined in great ape taxa to estimate numt insertion rate. Thirdly, alternative mechanisms of numt insertion, either through direct mitochondrial integration or post-insertional duplications, were also assessed. Fourthly, the effect of nuclear and mitochondrial environment on patterns of nucleotide composition and substitution was assessed through sequence comparisons of nuclear and mitochondrial paralogous sequences. Finally, numts in the gorilla genome were identified through two experimental methods and their use as polymorphic genetic markers was then evaluated in a sample of captive gorillas from U.S. zoos.
A deficit of MCR numts covering two particular mitochondrial subdomains was detected in all three apes examined, and is largely attributed to rapid loss of mitochondrial and nuclear sequence identity in the mitochondrial genome. Insertion rates have varied during the great ape evolution and exhibit substantial differences even between related taxa. The most likely mechanism of numt insertion is direct mitochondrial integration through Non-Homologous-End-Joining Repair. Transition/transversion ratios differed significantly between both mitochondrial and nuclear sequences and between numts from coding and non-coding mitochondrial regions. A previously documented upward bias in the GC content of the primate mitochondrial genome was confirmed and the extent of this bias relative to the corresponding numt sequences increased with numt age. Five gorilla-specific numts were isolated, including three exhibiting insertional polymorphisms that will be used in future population genetic studies in free-range gorilla.
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Lee, Sungkeun. "Molecular genetic analysis of nucleotide excision repair genes in Dictyostelium discoideum /." free to MU campus, to others for purchase, 1997. http://wwwlib.umi.com/cr/mo/fullcit?p9841209.
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Durewicz, Alicia Lynn. "Inheritance of chloroplast DNA (cpDNA) in Lobelia siphilitica." Kent State University Honors College / OhioLINK, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=ksuhonors1335812277.
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Pollock, Stephanie. "A study of genetic diversity and genome organization of Brassica napus using EST (expressed sequence tags) of Arabidopsis and SSR (simple sequence repeat) markers of B. napus /." Thesis, McGill University, 2001. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=33023.
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Arabidopsis expressed sequence tags (ESTs) and microsatellites of Brassica napus have been developed and used as PCR-based markers for both mapping and genetic diversity studies in B. napus . Out of 300 random Arabidopsis ESTs screened, 43 markers were mapped onto a genetic map of B. napus and then used in a diversity study involving 48 B. napus cultivars. A second set of EST markers were developed from chromosome 1 of Arabidopsis and used in genetic mapping studies of B. napus. From 192 primer pairs developed, 50 markers were added onto the B. napus reference map. Microsatellite markers were developed using a "GA" enriched genomic library from B. napus. From 152 designed primer pairs, 23 markers were added onto the B. napus reference map. Microsatellite markers were also used in genetic diversity studies of B. napus, where, from the 152 primer pairs, 40 revealed polymorphism between the 48 B. napus cultivars.
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Fobert, Pierre R. (Pierre Rheal) Carleton University Dissertation Biology. "Characterization of chromosomal sites of T-DNA integration by activation of a promoterless B-glucuronidase (GUS) gene linked to the T-DNA right border repeat." Ottawa, 1992.
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Conesa, Muñoz Miquel Àngel. "Hybridization patterns in Balearic endemic plants assessed by molecular and morphological markers." Doctoral thesis, Universitat de les Illes Balears, 2010. http://hdl.handle.net/10803/9373.
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La hibridació natural a plantes és un fenòmen àmplement conegut. És una important font de variabilitat que accelera l'evolució de les espècies. Es creu que és l'origen de moltes angiospermes, entre elles endemismes locals. Per altra banda, també pot tenir efectes negatius per la supervivència d'aquests endemismes, diluint els seus trets direfencials. En aquesta tesi s'estudia la possible hibridació natural que afecta a tres endemismes baleàrics (Viola jaubertiana, Lotus fulgurans i Helichrysum crassifolium), des del punt de vista dels marcadors moleculars basats en ADN i de la morfologia. S'avalua el paper de la hibridació natural la variabilitat, l'origen i la conservació d'aquestes espècies endèmiques.Natural hybridization is a widely known process in plants. It is an important source of variation promoting species evolution. It is likely to be the origin of many angiosperms, including local endemisms. Oppositely, it is also regarded as a potential threat for endemisms survivorship, diluting their differentail traits. This thesis deals with putative natural hybridization processes involving three Balearic endemics (Viola jaubertiana, Lotus fulgurans i Helichrysum crassifolium), from the points of view of the DNA molecular markers and the morphology. The role of natural hybridization in the variation, origin, and conservation of the above endemics is evaluated.
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Werkman, Joshua R. "DNA-BINDING SITE RECOGNITION BY bHLH AND MADS-DOMAIN TRANSCRIPTION FACTORS." UKnowledge, 2013. http://uknowledge.uky.edu/pss_etds/29.
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Herewithin, two transcription factor (TF) regulatory complexes were investigated. A bHLH–MYB–WDR (BMW) DNA-binding complex from maize was the first complex to be studied. R, a maize bHLH involved in the activation of genes in the anthocyanin pathway, had been characterized to indirectly bind DNA despite the presence of a functional DNA-binding domain. Findings presented here reveal that this is only partially correct. Direct DNA-binding by R was found to be dependent upon two distinct dimerization domains that function as a switch. This switch-like mechanism allows R to be repurposed for the activation of promoters of differing cis-element structure.
The second regulatory complex studied was of the Arabidopsis thaliana MIKC-MADS TF family. For many TFs, DNA-binding site recognition is relatively straightforward and very sequence specific, while others exhibit relaxed sequence specificity. MADS-domain TFs are one family of TFs with a wider range of cis-element sequences. Though consensus cis-element sequences have been determined for various MADS-domains, correctly predicting and identifying biologically functional cis-elements has been a challenge. In order to study the influence of nucleobase associations within the cis-element, a DNA-Protein Interaction (DPI)-ELISA method was modified and optimized to screen a panel of specific probes. Screening of the SEP3 homodimer against a panel of sequential, palindromic probes revealed that nucleobases in position -1:+1 of the CArG-box influence binding strength between the MADS-domain and DNA. Additionally, the specificity of AGL15 towards CT-W6-AG forms was discovered to be determined by the functional groups present in the minor groove at position -4:+4 using inosine:cytosine (I:C) base pairs.
Finally, the FLC–SVP MADS-domain heterodimer, bound to a native cis-element, was modeled and binding simulated using molecular dynamics. In conjunction with simulations of AGL15 and SEP3 homodimers, a potential binding mechanism was identified for this unique heterodimer. DNA sequence recognition by the MADS-domain was found to occur asymmetrically. In the case of the FLC–SVP heterodimer, the direction of asymmetrical DNA-binding in heterodimers was found to be fixed. Furthermore, the molecular dynamics simulations provided insight towards understanding the results generated from previous DPI-ELISA experiments, which should provide an improved means for predicting biologically significant CArG-boxes around genes.
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Boyko, Oleksandr, and University of Lethbridge Faculty of Arts and Science. "The versatile role of homologous recombination in plant cell : repair of DNA damage, stress-directed genome evolution and foreign DNA integration." Thesis, Lethbridge, Alta. : University of Lethbridge, Faculty of Arts and Science, 2008, 2008. http://hdl.handle.net/10133/724.
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Homologous recombination represents a DNA repair pathway. Its role in a plant cell is not limited to double strand break repair. It also extends to genome evolution via rearranging of DNA sequences, and has an important application in foreign DNA integration in the plant genome. Our study demonstrated that effects exerted by stress on homologous recombination and genome stability are not restricted to the exposed generation. The progeny of plants exposed to stress exhibited elevated spontaneous homologous recombination, changes in DNA methylation and higher tolerance to stress. These heritable changes are mediated by an unknown stress-inducible epigenetic signal. Furthermore, we demonstrated that using factors that enhance homologous recombination can improve the efficiency of genetic transformation by Agrobacterium. We have developed and patented a plant growth medium enhancing homologous recombination and significantly increasing the transformation frequency. The role of several other chemicals for the improvement of transformation was also evaluated.xxi, 246 leaves : ill. ; 29 cm. --
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Yu, Sung-Lim. "Analysis of the response of nucleotide excision repair genes in Dictyostelium discoideum /." free to MU campus, to others for purchase, 1997. http://wwwlib.umi.com/cr/mo/fullcit?p9841196.
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Munhoz, Carla de Freitas. "Diversidade genética de isolados de xanthomonas axonopodis pv. passiflorae com base em marcadores rep-PCR e AFLP e construção de primers específicos para diagnose." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/11/11137/tde-18052009-153241/.
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O patógeno Xanthomonas axonopodis pv. passiflorae causa a mancha oleosa ou bacteriose do maracujazeiro, uma doença que acarreta prejuízos à cultura em decorrência da baixa produção de frutos, podendo causar a morte das plantas. Uma coleção de 87 isolados deste patovar, oriundos de 22 cidades dos Estados de São Paulo, Minas Gerais e Paraná e do Distrito Federal, foi usada para estudar a diversidade genética por rep-PCR e AFLP. Nove isolados de outros patovares foram incluídos nas análises genéticas. A técnica rep-PCR revelou pouca diversidade entre os isolados do patovar passiflorae, mas diferenciou, claramente, os diferentes patovares. Todavia, a técnica AFLP revelou considerável diversidade genética entre os isolados do patovar passiflorae. A análise molecular da variância mostrou que a maior parte da diversidade (49,4%) se encontra entre as cidades de coleta. O agrupamento gerado com base nos coeficientes de similaridade e os resultados do teste de atribuição pelo programa Structure revelaram clusters genotípicos homogêneos. Isto evidencia que a variação está mais associada à geografia, ou seja, às cidades de coleta, e que o fluxo desses isolados é pequeno. Cinco conjuntos de primers foram desenhados para a detecção do patógeno em plantas. Desses, um conjunto de primers foi desenhado a partir da seqüência intergênica 16S-23S rRNA e se mostrou específico para o patovar passiflorae. Nenhum amplicon foi detectado nos patovares controles. O restante dos primers foi desenhado a partir do seqüenciamento de locos de AFLP, monomórficos para o patovar passiflorae e ausentes nos demais patovares. Estes primers não foram totalmente específicos; no entanto, todos podem ser recomendados para o diagnóstico da mancha oleosa, uma vez que não há registros de outras Xanthomonas em pomares de maracujá.The pathogen Xanthomonas axonopodis pv. passiflorae is responsible for the bacterial leaf spot of passion fruits, a disease that provokes commercial losses due to low levels of fruit production and even plant death. A group of 87 isolates of this pathovar, collected from 22 localities of São Paulo, Minas Gerais and Paraná States as in the Federal District was used to evaluate the genetic diversity based on rep-PCR and AFLP. Isolates from other nine pathovars were included in the genetic analyses. Low level of genetic diversity was revealed by the rep- PCR technique, which clearly distinguished the different pathovars. However, considerable diversity between isolates of the pathovar passiflorae was revealed by the AFLP technique. The analysis of molecular variance showed that differences between localities contributed to most part of the variance (49.4%). Groups generated based on similarity coefficients as well as results produced by the software Structure assigning isolates to groups, revealed homogeneous genotypic clusters. This confirms that variance is associated with geographic origin e.g. sampling localities, and that flow of isolates is restricted among localities. Five primer sets were designed for pathogen detection in plants; a primer set was designed for PCR amplification of the intergenic sequence 16-23S rRNA, which was shown to be specific to the pathovar passiflorae. No amplicons were detected in the controls. The remaining primers were designed after sequencing AFLP bands that were monomorphic within the pathovar passiflorae but absent in the other pathovars. These primers were not absolutely specific but all could be recommended for diagnosis of leaf spot as there is no report on the occurrence of other Xanthomonas species in passion fruit orchards.
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Iaffaldano, Brian. "Evaluating the Development and Potential Ecological Impact of Genetically Engineered Taraxacum kok-saghyz." The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1452174223.
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Coser, Sara Morra. "Diversidade em Psidium guajava L. por caracteres morfológicos, moleculares e citogenéticos." Universidade Federal do Espírito Santo, 2012. http://repositorio.ufes.br/handle/10/6635.
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Previous issue date: 2012-07-12A goiabeira (Psidium guajava L.) é uma das fruteiras de maior importância econômica da família Myrtaceae. O Brasil é um dos maiores produtores de goiaba do mundo, sendo esta uma cultura potencial em expansão e rentabilidade. A polinização cruzada da espécie e a existência de pomares heterogêneos de propagação seminal resultam em variabilidade, permitindo a seleção de genótipos para o melhoramento da cultura. O objetivo deste trabalho foi estudar a diversidade genética em genótipos de P. guajava selecionados em pomar de origem seminal e cultivares por características morfológicas e químicas de qualidade de fruto, também associar dados de conteúdo de DNA nuclear (2C), cariótipo, morfológicos e de marcadores moleculares microssatélites. Verificou-se a existência de divergência entre as Cortibel com relação às características de fruto, com genótipos apresentando performance superior e genótipos com desempenho semelhante à cultivares, potenciais para uso em hibridações ou como cultivares. As análises cariotípica e de conteúdo de DNA nuclear (2C) mostraram que os genótipos possuem um genoma estável, pequeno e diplóide e características cariotípicas relacionadas a grupos ancestrais de angiospermas. O dendrograma UPGMA baseado em dados morfológicos e SSR evidenciaram diversidade entre os genótipos, com melhor discriminação pelos dados de SSR. Como a maioria dos genótipos mostraram similaridade morfológica para as características de frutos, aliada a dissimilaridade molecular, estes se mostraram interessantes para o uso em hibridações em programas de melhoramento. O conjunto de dados gerados contribuiu para expandir o conhecimento sobre o genoma e a diversidade genética em P. guajava. Também é importante na estruturação de programas de melhoramento para a cultura, além de contribuir para estudos evolutivos
Guava (Psidium guajava L.) is one of the most economically important fruit crop from Myrtaceae family. Brazil is one of the largest producers of guava in the world, which is a potential crop in growth and profitability. Cross-pollination of the species and the existence of heterogeneous seminal propagation orchards result in variability, allowing the selection of genotypes for crop improvement. The aim of this study was to evaluate genetic diversity between P. guajava L genotypes selected from seminal origin orchard and cultivars, by morphological and fruit quality chemical characteristics, also associate data from nuclear 2C-value, karyotypic, morphological and simple sequence repeat (SSR) marker. There were divergences between Cortibel selections by fruit characteristics, with genotypes showing superior and similar performance when compared with cultivated genotypes, potential for use in hibridation and as cultivars. Karyotype and nuclear 2C-value analyses showed that all genotypes have a stable and very small diploid genome (2n = 2X = 22; 2C = 0.95 pg), and karyotypic characteristics related to ancestral angiosperm groups. UPGMA dendrogram based on morphological and SSR data evidenced diversity among the genotypes, with better discrimination by SSR data. Since most genotypes showed morphological similarity for fruit characteristics, combined with molecular dissimilarity, the use of these genotypes in hybridation breeding programs could be of interest. The obtained data set contributed to expand the knowledge about genome and genetic diversity of P. guajava. Also are important to structure crop improvement programs and contribute to evolutionary approaches
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Pereira, Renato Nunes. "Modelo hierárquico bayesiano na determinação de associação entre marcadores e QTL em uma população F2." Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/11/11134/tde-25042012-161429/.
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O objetivo do mapeamento de QTL (Quantitative Trait Loci ) e identificar sua posição no genoma, isto e, identificar em qual cromossomo esta e qual sua localização nesse cromossomo, bem como estimar seus efeitos genéticos. Uma vez que as localizações dos QTL não são conhecidas a priori, marcadores são usados frequentemente para auxiliar no seu mapeamento. Alguns marcadores podem estar altamente ligados a um ou mais QTL e, dessa forma eles podem mostrar uma alta associação com a característica fenotípica. O efeito genético do QTL e os valores fenotípicos de uma característica quantitativa são normalmente descritos por um modelo linear. Uma vez que as localizações dos QTL não são conhecidas a priori, marcadores são utilizados para representá-los. Em geral, e utilizado um numero grande de marcadores. Esses marcadores são utilizados no modelo linear para proceder ao processo de associação; dessa forma o modelo especificado contem um numero elevado de parâmetros a serem estimados. No entanto, e esperado que muitos destes parâmetros sejam não significativos, necessitando de um tratamento especial. Na estimação bayesiana esse problema e tratado por meio da estrutura de distribuições a priori utilizada. Um parâmetro que e esperado assumir o valor zero (não significativo) e naturalmente especificado por meio de uma distribuição que coloque um peso maior no zero, encolhimento bayesiano. Neste trabalho e proposta a utilização de dois modelos que utilizam distribuições a priori de encolhimento. Um dos modelos esta relacionado com o uso da distribuição a priori Laplace (Lasso bayesiano) e o outro com a Horseshoe (Estimador Horseshoe). Para avaliar o desempenho dos modelos na determinação da associação entre marcadores e QTL, realizou-se um estudo de simulação. Foi analisada a associação entre marcadores e QTL utilizando três características fenotípicas: produção de grãos, altura da espiga e altura da planta. Comparou-se os resultados obtidos neste trabalho com analises feitas na literatura na detecção dos marcadores associados a essas características. A implementação computacional dos algoritmos foi feita utilizando a linguagem C e executada no pacote estatístico R. O programa implementado na linguagem C e apresentado e disponibilizado. Devido a interação entre as linguagens de programação C e R, foi possível executar o programa no ambiente R.The objective of the mapping of quantitative trait loci (QTL) is to identify its position in the genome, ie, identify which chromosome is and what is its location in the chromosome, as well as to estimate their genetic eects. Since the location of QTL are not known a priori, markers are often used to assist in it mapping. Some markers may be closely linked to one or more QTL, and thus they may show a strong association with the phenotypic trait. The genetic eect of QTL and the phenotypic values of a quantitative trait are usually described by a linear model. Since the QTL locations are not known a priori, markers are used to represent them. Generally is used a large number of markers. These markers are used in the linear model to make the process of association and thus the model specied contains a large number of parameters to be estimated. However, it is expected that many of these parameters are not signicant, requiring a special treatment. In Bayesian estimation this problem is treated through structure priori distribution used. A parameter that is expected to assume the value zero (not signicant) is naturally specied by means of a distribution that put more weight at zero, bayesian shrinkage. This paper proposes the use of two models using priori distributions to shrinkage. One of the models is related to the use of priori distribution Laplace (bayesian Lasso) and the other with Horseshoe (Horseshoe Estimator). To evaluate the performance of the models to determine the association between markers and QTL, we performed a simulation study. We analyzed the association between markers and QTL using three phenotypic traits: grain yield, ear height and plant height. We compared the results obtained in this study with analyzes in the literature on the detection of markers associated with these characteristics. The computational implementation of the algorithms was done using the C language and executed the statistical package R. The program is implemented in C languages presented and made available. Due to the interaction between the programming languages C and R, it was possible execute the program in the environment R.
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Ostler, Jeffery Brent Jr. "Characterization of Pol IV and Pol V-Dependent Non-Coding RNAs Derived from aGeminivirus Genome." The Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1492698361649423.
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Raimundo, Luis Gustavo. "Homocisteína e cisteína séricas como marcadores epigenéticos de prognóstico e preditivos de resposta em tumores de mama." Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/5/5155/tde-06052014-125923/.
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O câncer de mama é a principal causa de mortalidade por câncer entre as mulheres. Alguns biomarcadores e características clínicas são utilizados para avaliar o prognóstico e prever a resposta a uma série de abordagens terapêuticas. A Homocisteína é conhecida como um fator de risco para doença vascular aterosclerótica, mas sua participação na biologia do câncer ainda é incerta. Cisteína é o aminoácido sulfurado derivado da Homocisteína no ciclo da Metionina. Este ciclo metabólico origina as bases nitrogenadas e também determina o nível de metilação da molécula de DNA. É atualmente reconhecido que a hipometilação global do genoma é um evento chave na transformação maligna das células. O objetivo deste estudo foi avaliar os níveis séricos de homocisteína e cisteína como biomarcadores de sobrevida e de progressão da doença em câncer de mama. Também foi avaliado o efeito de um curso de curta duração (um mês) de tratamento hormonal sobre os níveis de Homocisteína, Cisteína e metilação do DNA. Amostras de sangue foram obtidos por ocasião da biópsia inicial (pré-tratamento) em todas as pacientes e, de tumor e de tecido normal adjacente, ao diagnóstico eem um mês após, para as pacientes que receberam o regime hormonal neo-adjuvante (pré-operatório). Todas as pacientes eram mulheres na pós-menopausa, com tumores de mama ressecáveis, acompanhadas em dois hospitais públicos, que consentiram em participar de outros dois protocolos de pesquisa prévios. Homocisteína e Cisteína foram analisadas por HPLC e a metilação global do DNA do tecido foi determinada por meio da técnica de MSRE (Methylation-Sensitive Restriction Enzyme). Foi observada uma diferença significativa entre os níveis pré e póstratamento de Homocisteína e Cisteína em tumores avançados, sugerindo um papel prognóstico em pacientes com características clínicas reservadas. As variações nos níveis de Homocisteína se mostraram significativamente correlacionadas com a sobrevida livre de doença. O modelo de risco proporcional de Cox demonstrou que os níveis de homocisteína e o status dos linfonodos representaram fatores prognósticos independentes em termos de sobrevida livre de doença. Embora mais estudos sejam necessários para confirmar estes resultados, nossa pesquisa sugere que a Homocisteína pode ser usada como um biomarcador de prognóstico para câncer de mamaBreast cancer is the leading cause of cancer mortality among women. Some biomarkers and clinical features are used to evaluate prognosis and to predict response to a range of therapeutic approaches. Homocysteine is well known as a risk factor in atherosclerotic vascular diseases, but its participation in cancer biology is still unclear. Cysteine is a sulfur amino acid derived from Homocysteine in the Methionine cycle. This metabolic cycle originates the nitrogenous bases and determines the methylation level of the DNA molecule as well. It is currently recognized that the global hipomethylation of the genome is a key event in the malign transformation of cells. The aim of this study was to evaluate serum Homocysteine and Cysteine as biomarkers of survival and disease progression in breast tumor, as well as the methylation status of tumor and normal tissues. The effect of a short course (one month) of hormonal treatment on Homocysteine, Cysteine and DNA methylation levels was also evaluated. Blood samples were collected during the initial biopsy (pretreatment) in all patients and, tumor samples and normal adjacent tissue, at diagnosis and one month after, for the patients that received neo-adjuvant hormonal regimen (pre-treatment). All patients were post-menopausal women, with resectable breast tumors, followed at two public hospitals, and that had consented to participate in two previous research protocols related to their disease. Serum Homocysteine and Cysteine were analyzed by HPLC and tissue global DNA methylation was determined by the MSRE (Methylation- Sensitive Restriction Enzyme) technique. A significant difference was observed between pre- and post-treatment levels of Homocysteine and Cysteine in advanced tumors, suggesting a prognostic role in patients with poor clinical characteristics. Variations in Homocysteine levels were significantly correlated with disease free survival. Cox proportional risk model demonstrated that nodal status and Homocysteine levels were independent prognostic factors for Disease Free Survival. Although more studies are needed to confirm these results, our research suggests that Homocysteine might be used as a prognostic biomarker for breast cancer
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Boudko, Ekaterina. "Phylogenetic Analysis of Subtribe Alopecurinae (Poaceae)." Thèse, Université d'Ottawa / University of Ottawa, 2014. http://hdl.handle.net/10393/30696.
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Subtribe Alopecurinae (Poeae, Poaceae) sensu lato‘s seven genera share interesting morphological similarities (dense spicate panicles and one-flowered spikelets) that were widely thought to have a common origin. However, recent molecular evidence for three of the genera has suggested that the subtribe may be polyphyletic. To test this, five DNA regions were sequenced and analyzed using phylogenetic methods. Results confirm that Alopecurinae s.l. as presently treated is polyphyletic and should be dissolved. Additionally, the genus Cornucopiae may be just another Alopecurus. Limnas and Pseudophleum are not closely allied to Alopecurus or each other, and are even further from Phleum. Phleum is a distinct lineage that is not closely allied to any other included Alopecurinae genus. Evidence for revising infrageneric classifications of Alopecurus and Phleum is presented, as is evidence for separating A. magellanicus into two or more subspecies.
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Erasmus, Tertia Elizabeth. "Genetic diversity of proprietary inbred lines of sunflower, determined by mapped SSR markers and total protein analysis." Thesis, 2008. http://hdl.handle.net/10413/879.
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Hongtrakul, Vipa. "The development and analysis of sequence-based DNA markers in sunflower for DNA fingerprinting and candidate gene analysis." Thesis, 1997. http://hdl.handle.net/1957/33809.
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Molecular DNA markers have become widely used in all areas of genetic
research. The objectives of this thesis were to develop polymorphic markers in sunflower
and utilize the markers for genetic and candidate gene analyses. Amplified fragment
length polymorphism (AFLP) markers were used to estimate genetic similarities and
assess the genetic diversity among 24 public oilseed inbred lines of sunflower
(Helianthus annuus L.). A total of 359 AFLP markers were scored by using six AFLP
primer combinations. Genetic similarities ranged from 0.70 to 0.91, polymorphism rate
ranged from 7 to 24%, and polymorphic information contents (PICs) ranged from 0.0 to
0.5. Principal coordinate and cluster analysis separated the lines into two groups, B-lines
and R-lines, illustrating breeding history, basic heterotic pattern and the widespread
practice of using each group to develop new lines.
��9 stearoyl-ACP desaturase (SAD) and ��l2 oleate desaturase (OLD) cDNAs
were cloned and sequenced. DNA fragment length polymorphism (DFLP), single strand
conformational polymorphism (SSCP), and simple sequence repeat (SSR) markers were
developed for the SAD6 and SAD17 genes among eight elite inbred lines. PICs for DFLP, SSCP, and SSR markers were 0.18, 0.37, and 0.30, respectively. Length variants were due to long monomeric repeats, insertions, and deletions in intron sequences, thereby producing polymorphic markers.
OLD desaturates 18:1-PC (oleoyl phosphatidylcholine) to 18:2-PC, thereby converting oleic to linoleic acid. It is a likely candidate gene to be causing the high oleic phenotype in mutant sunflower. The expression of OLD7 in developing seeds was greatly reduced in mutant as opposed to wildtype backcross-derived lines. The restriction fragment length polymorphism (RFLP) patterns suggest that OLD7 is duplicated and rearranged in mutant lines.
Utilizing sunflower SAD gene sequences and 27 inbred lines, intron fragment length polymorphism (IFLP) markers were developed for automated genotyping. These IFLP markers with ~470 to ~850 bp in length had a mean PIC score of 0.414, versus 0.336 for DFLP markers, and 0.582 for SSCP markers. One and two nucleotide length polymorphisms were reliably detected in PCR fragments up to ~150 and ~680 bp, respectively.Graduation date: 1998
35
Nsabimana, Antoine. "Establishing genetic diversity of Rwanda highland banana using random amplified polymorphic DNA markers." Thesis, 2006. http://hdl.handle.net/10413/4670.
Full textAbstract:
The characterization of the banana germplasm collection from Rubona - Rwanda was
investigated using morphological and cytological characteristics of the genomic groups.
Genetic diversity was assessed using Random Amplified Polymorphic DNA analysis.
The survey was conducted to evaluate the distribution of banana cultivars in the four
major growing regions of Rwanda.
A total of 90 accessions from the National Banana Germplasm Collection at Rubona
Rwanda were characterized and six characters of the fingers (length, width, weight,
green life, post green life and length/width ratio) were subjected to principal component
analysis (PCA). The cooking and beer clones were separated. The cooking clones were
further grouped into three clone sets: Musakala, Nakabululu, and one that constitutes
Nakitembe and Nfuuka clone sets. The AAB genomic group was separated from AAA,
AB and ABB genomic groups.
The results from the survey showed that East African Highland bananas are the most
important genotype group in the four major banana growing regions of Rwanda ranging
between 60 - 90% of banana mats counted. Several new Highland banana cultivars
were recorded, such as 'Intokatoke', 'Igihuna', 'Ingenge', 'Ingaju', 'Icyerwa', 'Mitoki',
'Madamu', 'Inkokobora', 'Intokekazi', 'Bugoyi', 'Ishoki'. Amongst these cultivars, some
were classified as cooking and others as brewing bananas. However, in the National
Banana Germplasm Collection at Rubona - Rwanda, the uses of these cultivars are
recorded differently therefore increasing the need for agro-morphological
characterization.
The assessment of ploidy level of accessions from the National Banana Germplasm
Collection at Rubona - Rwanda, by flow cytometry showed misclassification of some
accessions such as 'Pomme', 'Kamaramasenge', 'Gisubi kayinja', 'Gisubi kagongo', and
'Dibis' which were classified as diploid, diploid, triploid, and tetraploid respectively. They
IV
were found to be triploid, triploid, triploid, diploid and triploid. All these bananas were
recently introduced into Rwanda, while the endemic Highland bananas were triploid.
The genomic group and genetic similarities of 49 accessions were investigated using
Random Amplified Polymorphic DNA markers. The genomic group of bananas
assessed were established using OPA-18 (PILLAY et al., 2000) and OPG-17 primers.
These primers showed bands 441 and 443 base pairs (bp) respectively for the
accessions having only the B genome. Whilst they were absent for the accessions
"
having an A genome. The genetic similarity was estimated via a Simple Matching
coefficient which showed the lowest value 0.46 measured between 'Ingumba' and
'Ishika 'and the highest value of 0.85 between 'Kirayenda' and 'Inyabukuwe'. The data
of matrix of coefficient of similarity was subjected to cluster analysis with unweighted
pair group method with arithmetic average (UPGMA). Each accession was clearly
separated demonstrating the usefulness of RAPDs in analysis of genetic diversity. The
results of this study are very important to the Curator of the banana germplasm
collection in Eastern Central Africa and for the future breeding of this crop.Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2006.
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Marian, Calin O. Bass Hank W. "Identification of maize (Zea mays L.) genes encoding telomere repeat DNA-binding proteins." Diss., 2005. http://etd.lib.fsu.edu/theses/available/etd-07082005-173029.
Full textAbstract:
Thesis (Ph. D.)--Florida State University, 2005.Advisor: Dr. Hank W. Bass, Florida State University, College of Arts and Sciences, Dept. of Biological Science. Title and description from dissertation home page (viewed Sept. 19, 2005). Document formatted into pages; contains viii, 93 pages. Includes bibliographical references.
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Thomas, Stephen W. (Stephen William). "Molecular studies of homologous chromosome pairing in Triticum aestivum." 1997. http://web4.library.adelaide.edu.au/theses/09PH/09pht462.pdf.
Full textAbstract:
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.
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Msomi, Nhlanhla Sobantu. "The potential of bulk segregant analysis and RAPD technology for identification of molecular markers linked to traits in sugarcane." Thesis, 1998. http://hdl.handle.net/10413/4403.
Full textAbstract:
The objective of the present study was to investigate the potential use of bulk segregant analysis (Michelmore et al., 1991) as a method to rapidly identify genetic markers linked to traits in sugarcane. Four bulked DNA samples were prepared from progeny of a sugarcane cross, AA157, based on segregation for the fibre trait. The bulks comprised five and ten individuals on either side of the fibre phenotypic extreme. The random amplified polymorphic DNA (RAPD) technique (Williams et ai., 1990) was used to screen for differences between the low and high fibre bulks. A total of 749 fragments were amplified in the bulks, eight of which were polymorphic. The segregation of the bulk specific polymorphism was analysed in 80 progeny of the AA157 cross; and seven were found to reproducibly segregate on a 1: 1 basis. This indicates that they are single dose fragments. A total of 79 polymorphisms were detected between the parents of the cross, indicating 10.5% variation in the genomic region sampled. Twenty two of the parental polymorphisms segregated as single dose fragments in the progeny of the cross AA157. Analyses of variance (ANOVAs), and multiple regression analyses, were used to ascertain linkage of the putative RAPD markers to fibre, and if linked, to determine the fibre variation ascribed respectively. Three RAPD fragments were found linked to the fibre trait. Fragments OPA17438 and OPC16889 (at the 99% significance level), and OPB1l464 (at the 95% significance level). These putative markers ascribed a total of 28.6% fibre variation in the 1993 season. The association of the RAPD markers with fibre in the different seasons (1992, 1993 and 1994) was investigated. Three RAPD markers were found linked to the fibre trait in each season, with a total of 5.5% and 31,4% fibre variation ascribed in the 1992 and 1994 seasons respectively. Marker OPA17438 was found to be linked to the fibre trait in all three seasons investigated, and marker OPC16889, was found linked to the fibre trait in the 1992 and 1993 seasons. Cross validation of the linkages of the RAPD markers to the fibre trait was carried out by a modified form of 'jacknifing' where the sample size was reduced to N-l0, and RAPD marker-fibre trait associations investigated as before. RAPD markers OPA17438 and OPC16889 were still consistent across the seasons, however marker OPA17438 was no longer linked to the fibre trait in the 1992 season. To investigate the genetic behaviour of RAPD based markers in sugarcane and the potential for their application in marker-assisted selection (MAS), two putative RAPD markers were converted to sequence characterised amplified regions (SCARs) (Paran and Michelmore, 1993). The RAPD fragments OPA17438, OPBl1464, and OPC16889 were excised from agarose gels, re-amplified and cloned into the pCR-Script SK (+) phagemid for sequencing. RAPD markers OPA17438 and OPB11 464 were converted to SCARs by using their sequences to design longer specific primers. A third SCAR marker, SAl1640, originally derived from sugarcane cDNA as a potential stem preferential expressed sequence tag, was included in the analysis to increase the sample size. All three SCAR markers segregated in a monomorphic fashion in the parents and progeny of the cross AA157. In addition, monomorphic length variants for markers, OPA17438 and OPB11 464 were detected with the SCAR amplification. All three SCARs segregated in a monomorphic fashion in different commercial varieties and bulks of S. officinarum and S. spontaneum, the progenitors of modern commercial varieties. The segregation analyses of the SCAR markers indicate that the RAPD polymorphism of marker SAl1640 was probably due to a point mutation or mismatch in the priming site. The segregation analyses of SCARs for the markers OPA17438 and OPB11464 indicate that their segregation in the RAPD analyses was due to an insertion mutation in the genetic locus. The combined results of the SCAR and RAPD segregation of markers OPA17438 and OPB11464 are indicative of preferential pairing in the cross AA157. Finally, to investigate the extent of linkage disequilibrium in a modern commercial variety, twenty two single dose RAPD fragments were investigated for their association with four traits in 53 progeny of cross AA157. The four traits investigated were fibre %cane, brix %cane, pol %cane and ers %cane over three seasons (1992, 1993 and 1994), at different ages of harvest (12, 8, and 9 months respectively). Seventeen linkages of RAPD markers to the four traits, over the three seasons, were detected. The phenotypic variation ascribed by the RAPD markers ranged from 7.6% fibre %cane variation explained by one marker in 1992, 29.6% fibre %cane (three markers) in the 1993 season to 10% (three markers) in 1994. A total of 14.1% brix %cane variation was ascribed by two markers in 1992, 9.6% (one marker) in 1993 and 16.3% (two markers) in the 1994 season. A total of 13.5% estimated recoverable sucrose %cane was ascribed by one marker in 1992, 12% (two markers) in 1993 and 15.3% (two markers) in the 1994 season. Two markers explained 17.2% pol %cane variation in 1992 and 25.4% in the 1994 season. Only four markers were detected across different environments, three of which were linked to fibre. These were OPA17438, OPB16618 and OPC16889, each linked to fibre in two seasons. RAPD marker OPB11 464 was linked to estimated recoverable sucrose %cane in two seasons. Two markers were found associated with different traits in a single season. RAPD marker OPB11 464 was found associated with brix %cane and estimated recoverable sucrose %cane in the 1993 season, and RAPD marker OPA17438 was found associated with all four traits in the 1994 season.Thesis (Ph.D.)-University of Natal, Durban, 1998.
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Taylor, Christopher 1966. "Cytogenetic and molecular genetic markers for chromosome 6R of rye linked to CCN resistance / by Christopher Taylor." 1996. http://hdl.handle.net/2440/18939.
Full textAbstract:
Includes bibliographies.xiv, 175, [96] leaves, [17] leaves of plates : ill. (some col.) ; 30 cm.
Title page, contents and abstract only. The complete thesis in print form is available from the University Library.
This thesis reports on the generation of molecular tools for the analysis of chromosome 6R of rye and the application of these tools in structural analysis of 6RL. Results presented include physical and genetic maps of chromosome 6RL incorporating RFLP and PCR markers and CreR, the locus conferring resistance to cereal cyst nematode (CCN). The ability to detect small introgessions of rye chromatin in wheat is demonstrated.
Thesis (Ph.D.)--University of Adelaide, Dept. of Plant Science, 1997
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"Identification and characterization of differentially expressed genes in dikaryons of lentinula edodes by cDNA microarray." 2004. http://library.cuhk.edu.hk/record=b5896199.
Full textAbstract:
by Shih Sheung Mei.Thesis submitted in: July 2003.
Thesis (M.Phil.)--Chinese University of Hong Kong, 2004.
Includes bibliographical references (leaves 206-215).
Abstracts in English and Chinese.
Abstract --- p.ii
Achnoledgements --- p.vi
Abbreviations --- p.viii
List of contents --- p.viv
List of tables --- p.xiii
List of figures --- p.xv
Chapter Chapter One --- Literature Review
Chapter 1.1 --- Introducation of Lentinula edodes --- p.1
Chapter 1.1.1 --- Life cycle of Basidiomycete --- p.1
Chapter 1.1.2 --- Differentially Expressed Genes in stages of Lentinula edodes --- p.3
Chapter 1.2 --- Relationship of Monokaryons and Dikaryons in Basidiomycetes --- p.4
Chapter 1.2.1 --- Mating Type Gene in Filamentous Fungi --- p.4
Chapter 1.2.3 --- Dikaryon Formation and Homeodomain Proteins --- p.6
Chapter 1.2.4 --- Clamp Connection formation in Dikaryon --- p.9
Chapter 1.3 --- Stuctural Protein of Mushroom --- p.11
Chapter 1.3.1 --- Hydrophobin --- p.11
Chapter 1.3.1.1 --- General Introduction --- p.11
Chapter 1.3.1.2 --- Structure of hydrophobin --- p.11
Chapter 1.3.1.3 --- Formation of Disulphide bonds and Glycosylation --- p.12
Chapter 1.3.1.4 --- Functions of Hydrophobins --- p.13
Chapter 1.4 --- Genomics of filamentous fungi --- p.15
Chapter 1.5 --- Genetic analysis of filamentous fungi --- p.18
Chapter 1.6 --- Objectives of the Project --- p.20
Chapter Chapter Two --- Identification of Differentially Expressed Genes in Dikaryons of Lentinula edodes by Microarray of Primordium Expressed Sequence Tags
Chapter 2.1 --- Introduction --- p.23
Chapter 2.2 --- Materials and Methods --- p.27
Chapter 2.2.1 --- Construction of EST database --- p.27
Chapter 2.2.2 --- Construction of EST Microarray cDNA gene-chip --- p.27
Chapter 2.2.2.1 --- Amplification of the primordium EST clones --- p.27
Chapter 2.2.2.2 --- Purification of the amplified EST clones --- p.28
Chapter 2.2.2.3 --- Spotting of the amplified EST clones onto chips --- p.29
Chapter 2.2.3 --- Screening of the Differentially Expressed Genes in Dikaryons by Primordium Microarray --- p.31
Chapter 2.2.3.1 --- Mycelium Cultivation and Preparation of Total RNA --- p.31
Chapter 2.2.3.2 --- cDNA synthesis and labeling --- p.32
Chapter 2.2.3.3 --- cDNA purification --- p.33
Chapter 2.2.3.4 --- Probe Storage Conditions --- p.34
Chapter 2.2.3.5 --- cDNA analysis --- p.35
Chapter 2.2.3.6 --- Microarray hybridization --- p.37
Chapter 2.2.3.7 --- Stringency washes --- p.39
Chapter 2.2.3.8 --- Detection with TSA --- p.39
Chapter 2.2.3.9 --- Microarray scanning and data anlysis --- p.41
Chapter 2.3 --- Results --- p.45
Chapter 2.3.1 --- Amplification of primordium ESTs --- p.45
Chapter 2.3.2 --- Purification of PCR products --- p.45
Chapter 2.3.3 --- Data Analysis of Microarray Data --- p.47
Chapter 2.3.3.1 --- Generation of Primordium EST Microarray Image for analysis --- p.47
Chapter 2.3.3.2 --- Normalization of the Data --- p.49
Chapter 2.3.3.3. --- Transciption Profile of Dikaryon compared with Monokaryon --- p.79
Chapter 2.3.3.4. --- Differentially Expression of Dikaryon L54 --- p.80
Chapter 2.4 --- Discussion --- p.85
Chapter Chapter Three --- Enrichment of Genes with Differentially Expression in Dikaryons by Construction of Full-length Subtractive Library
Chapter 3.1 --- Introduction of Subtraction Cloning --- p.93
Chapter 3.2 --- Materials and Methods --- p.97
Chapter 3.2.1 --- Construction of Full-length Dikaryotic Subtractive library --- p.97
Chapter 3.2.1.1 --- Isolation of PolyA+ mRNA of Dikaryon for Subtraction --- p.97
Chapter 3.2.1.2 --- Enrichment of Differentially Expressed Genes in Dikaryon L54 by Subtraction with Monokaryons A and B --- p.99
Chapter 3.2.1.3 --- First-Strand cDNA Synthesis --- p.102
Chapter 3.2.1.4 --- cDNA Amplification by Long-Distance PCR --- p.102
Chapter 3.2.1.5 --- Proteinase K Digestion --- p.103
Chapter 3.2.1.6 --- Sfi Digestion --- p.104
Chapter 3.2.1.7 --- cDNA size fractionation by CHROMA SPIN-400 --- p.104
Chapter 3.2.1.8 --- Determination of the Ligation Efficiency --- p.106
Chapter 3.2.1.9 --- Ligation of cDNA to lamda TriplEx2 Vector --- p.107
Chapter 3.2.1.10 --- Lamda-phage Packaging Reaction --- p.107
Chapter 3.2.1.11 --- Titering the Unamplifled Library and Determining the Percentage of Recombinant Clones --- p.108
Chapter 3.2.1.12 --- Library Amplification --- p.109
Chapter 3.2.1.13 --- Conversion of λTriplEx2 Recombinant Clones to pTriplEx2 Recombinant Plasmids --- p.111
Chapter 3.2.2 --- Screening of the Subtractive library --- p.114
Chapter 3.2.2.1 --- Verification of the enrichment by Plaque Lifting hybridization --- p.114
Chapter 3.2.2.1.1 --- Lifting the Plaques --- p.114
Chapter 3.2.2.1.2 --- Synthesis of the Probes for Plaque Lift Hybridization --- p.115
Chapter 3.2.2.1.3 --- Hybridization to the Membranes --- p.116
Chapter 3.2.2.2 --- Screening the Subtractive library by Macroarray Hybridization --- p.117
Chapter 3.2.2.2.1 --- Colony Picking by QPik System --- p.117
Chapter 3.2.2.2.2 --- Gridding of Macroarray --- p.118
Chapter 3.2.2.2.3 --- Filter Processing of Gridded Membrane --- p.119
Chapter 3.2.2.2.4 --- Hybridization to the Macroarray Membrane --- p.120
Chapter 3.3 --- Results and Discussion --- p.121
Chapter 3.3.1 --- Enrichment of Differentially Expressed Genes in Dikaryon L54 by Subtraction with Monokaryons A and B --- p.121
Chapter 3.3.2 --- Construction of the full-length subtractive library --- p.123
Chapter 3.3.3 --- Conversion of A TriplEx2 Recombinant Clones to pTriplEx2 Recombinant Plamid --- p.124
Chapter 3.3.4 --- Verification the Enrichment of Subtractive library by Plaque lifting Hybridization --- p.125
Chapter 3.3.5 --- Screening of the Subtractive library by Macroarray --- p.125
Chapter 3.4 --- Discussion --- p.126
Chapter Chapter Four --- Identification of Genes with Differentially Expression in Dikaryons by Subtactive cDNA Library Microarray
Chapter 4.1 --- Introduction --- p.135
Chapter 4.2 --- Materials and Methods
Chapter 4.2.1 --- Selection and Amplification of clonesin SubtractionLlibrary for Microarray screening --- p.140
Chapter 4.2.2 --- PCR product Purification --- p.141
Chapter 4.2.3 --- Generation of Subtractive Dikaryotic Library Microarray Chip --- p.142
Chapter 4.2.4 --- Screening the Differentially Expressed Genesin Dikaryon L54 by the Subtraction Dikaryotic Library cDNA Microarray Analysis --- p.143
Chapter 4.2.4.1 --- Preparation of Total RNA --- p.143
Chapter 4.2.4.2 --- Synthesis and fluorescent labeling of total cDNA --- p.145
Chapter 4.2.4.3 --- Purification of labeled cDNA --- p.146
Chapter 4.2.4.4 --- Storage Condition of Probe --- p.147
Chapter 4.2.4.5 --- Analysis of labeled total cDNA --- p.148
Chapter 4.2.4.6 --- Microarray hybridization --- p.150
Chapter 4.2.4.7 --- Stringency washes --- p.152
Chapter 4.2.4.8 --- Detection with TSA --- p.153
Chapter 4.2.4.9 --- Image generation and data analysis --- p.155
Chapter 4.2.5 --- Sequence analysis of clones showing differentially expressed in dikaryons in microarray screening --- p.157
Chapter 4.2.5.1 --- Single-pass partial sequencing of 3´ة-end of subtractive cDNA clones --- p.157
Chapter 4.2.5.2 --- Compiling dikaryotic EST database --- p.158
Chapter 4.2.6 --- Comparison microarray analysis with SAGE analysis of the differentially expressed genes --- p.159
Chapter 4.3 --- Results --- p.161
Chapter 4.3.1 --- Preparation of clones for microarray hybridization --- p.161
Chapter 4.3.2 --- Screening the differentially expressed genesin dikaryon L54 by the subtractive dikaryotic library cDNA microarray analysis --- p.162
Chapter 4.3.2.1 --- Image capture and microarray data analysis --- p.162
Chapter 4.3.2.2 --- Comparision of dikaryon L54 with monokaryons A and B --- p.163
Chapter 4.3.2.3 --- Sequenced and comparison of the differentially expressed genes in dikaryon --- p.166
Chapter 4.3.3 --- Comparison microarray analysis with SAGE analysis of the differentially expressed genes --- p.169
Chapter Chapter Five --- Conclusion and Future Perpectives --- p.198
References --- p.206
41
Wilcox, Buck W. L. "Effects of DNA mismatch repair inhibition in Arabidopsis thaliana." Thesis, 2012. http://hdl.handle.net/1957/28606.
Full textAbstract:
Genomic instability underlies diseases of unregulated cell growth that result in
cancers and developmental abnormalities in humans. Similar genome destabilizing
mechanisms are used to create genetic variety in crops for use in breeding and trait
development. Errors that occur during DNA replication may cause mutations if
they are not corrected before further cell divisions. DNA mismatch repair
(MMR) corrects misinsertions and insertion/deletion DNA loop-outs that arise
during DNA replication in plants, animals, prokaryotes, and some archaea, all of
which incur mutations at rates 100 to 1,000-fold greater when subjected to
inherited or somatic-mismatch repair deficiencies. An understanding of the
effects of mismatch repair on somatic and germ-line cells in Arabidopsis thaliana is
critical to the development of this plant as a model system for the study of
genomic instability. Insertions and deletions of multiples of two base pairs in
dinucleotide repeat sequences (microsatellites) occur more frequently in the
absence of mismatch repair, and the mismatch-repair status of an individual,
tissue, or cell may be inferred on the basis of microsatellite mutation frequency.
Single-template PCR analysis measured microsatellite mutation frequencies in
leaves and shoot-apical-meristem stem cells, and allowed me to address for the
first time an important question: Do plants relax mismatch repair in vegetative
tissues relative to meristematic germ-line and floral tissue? Analyses of four
microsatellite loci in mismatch repair-deficient and wild type plants surprisingly
suggest that there is little difference in mismatch repair activity between leaves and
seeds. Mismatch-repair-deficient leaves displayed only two-fold higher
microsatellite mutation frequency compared to wild type, and wild-type leaves also
displayed a two-fold higher microsatellite mutation frequency compared to shoot-apical-
meristems. The high frequency of microsatellite mutation in these wildtype
tissues is unexpected, and it suggests that plants relax mismatch repair in
differentiated tissues while maintaining genetic fidelity in a small set of stem cells
in the shoot apical meristem (SAM). Genome sequencing of msh2⁻/⁻ mutation
accumulation A. thaliana lines provides an estimated germ-line mutation rate of
3.9 × 10⁻⁷ in the absence of mismatch repair. Comparison of the rates of base
substitution mutation per chromosome in mismatch repair-deficient plants with
rates reported for wild-type plants suggests mismatch repair is more efficient on
chromosome 5 than on chromosomes 1-4. Bias towards G:C → A:T mutations
among transitions is maintained but increased nearly 100-fold in the absence of
mismatch repair.Graduation date: 2012
42
Peterschmidt, Brooke C. "DNA markers and characterization of novel sources of eastern filbert blight resistance in European hazelnut (Corylus avellana L.)." Thesis, 2013. http://hdl.handle.net/1957/37973.
Full textAbstract:
European hazelnut is a significant crop in the Pacific Northwest, and the US ranks
4th internationally for hazelnut production. Production in the Pacific Northwest is
threatened, however, by the disease eastern filbert blight (EFB) caused by the fungus
Anisogramma anomala (Peck) E. Müller. To meet the challenges faced by the hazelnut
industry in Oregon and Washington, the breeding program at Oregon State University has
focused on developing DNA marker technology and producing EFB resistant cultivars.
This study focused on developing new microsatellite markers from hazelnut
transcriptome sequences and on disease resistance from three accessions ('Culpla,' 'Crvenje,' and OSU 495.072) which showed no disease symptoms following a series of
inoculations.
DNA markers have been useful in hazelnut breeding for marker-assisted
selection, construction of genetic linkage maps, cultivar fingerprinting, and phylogeny
studies. Previously developed markers include AFLP, RAPD, ISSR, and microsatellite
(SSR) markers developed from enriched libraries and ISSR fragments. This study utilized
the transcriptome sequence from 'Jefferson' hazelnut to mine for microsatellites, align
with the genomic sequence, design primers, screen for polymorphism, and characterize
and map polymorphic markers. A total of 1432 microsatellites were mined from the
transcriptome sequence, and the most frequently found motifs were AG (35.8%), AT
(13.3%), and AAG (12.7%), and 382 primer pairs were designed. Screening showed that 119 markers were polymorphic, and these were characterized on sets of 50 and 14 accessions. Fifty-three markers that segregated in the mapping population or in three alternate populations were mapped and assigned to linkage groups. A dendrogram showed that accessions clustered mostly according to geographic origin. These results confirm the high level of diversity present in hazelnut, and the markers developed in this study will be useful for further genetics studies in hazelnut.
The three EFB resistant parents 'Culpla,' 'Crvenje,' and OSU 495.072 were subjected to two inoculation treatments: greenhouse inoculations and exposure under an inoculation structure. The accessions remained free of disease after both treatments. Progeny segregating for resistance were produced. The progeny were inoculated either in the greenhouse or under the structure, and disease response recorded for each individual. DNA was extracted from seedlings, and sets of 32 seedlings from each resistant parent were screened with previously mapped markers using PCR and capillary electrophoresis. All three resistance sources were correlated with marker A614, allowing the resistance loci to be assigned to linkage group (LG) 6. The progeny were then screened with all known microsatellite markers on LG 6, and linkage maps constructed of the marker loci and resistance loci. Markers KG821, LG628, and LG696 are especially close to the resistance loci and will be useful for marker-assisted selection. Although these resistance loci are located in the same region of LG 6 as the 'Gasaway' resistance gene, they are different from 'Gasaway,' and markers linked to resistance will be useful for introgressing and pyramiding resistance in new cultivars.Graduation date: 2013
43
Kögler, Anja. "Diversity and Evolution of Short Interspersed Nuclear Elements (SINEs) in Angiosperm and Gymnosperm Species and their Application as molecular Markers for Genotyping." 2019. https://tud.qucosa.de/id/qucosa%3A72118.
Full textAbstract:
Short interspersed nuclear elements (SINEs) are small non-autonomous and heterogeneous retrotransposons, widespread in animals and plants and usually differentially propagated in related species resulting in genome-specific copy numbers.
Within the monocots, the Poaceae (sweet grasses) is the largest and economically most important plant family. The distribution of 24 Poaceae SINE (PoaS) families, five of which showing a subfamily structure, was analyzed in five important cereals (Oryza sativa, Triticum aestivum, Hordeum vulgare, Sorghum bicolor, Zea mays), the energy crop Panicum virgatum and the model grass Brachypodium distachyon. The comparative investigation of SINE abundance and sequence diversity within Poaceae species provides insights into their species‐specific diversification and amplification. The PoaS families and subfamilies fall into two length and structural categories: simple SINEs of up to 180 bp and dimeric SINEs larger than 240 bp. Of 24 PoaS families, 20 are structurally related across species, in particular either in their 5′ or 3′ regions. Hence, reshuffling between SINEs, likely caused by nested insertions of full-lengh and truncated copies, is an important evolutionary mechanism of SINE formation. Most striking, the recently evolved homodimeric SINE family PoaS‐XIV occurs exclusively in wheat (T. aestivum) and consists of two tandemly arranged PoaS‐X.1 copies.
Exemplary for deciduous tree species, the evolutionary history of SINE populations was examined in six Salicaceae genomes (Populus deltoides, Populus euphratica, Populus tremula, Populus tremuloides, Populus trichocarpa, Salix purpurea). Four of eleven Salicaceae SINE (SaliS) families exhibit a subfamily organization. The SaliS families consist of two groups, differing in their phylogenetic distribution pattern, sequence similarity and 3’ end structure. These groups probably emerged at different evolutionary periods of time: during the ‘salicoid duplication’ (~ 65 million years ago) in the Salix-Populus progenitor, and during the separation of the genus Salix (~ 45 - 65 million years ago), respectively. Similar to the PoaS families, the majority of the 20 SaliS families and subfamilies share regions of sequence similarity, providing evidence for SINE emergence by reshuffling. Furthermore, they also contain an evolutionarily young dimeric SINE family (SaliS-V), amplified only in two poplar genomes. The special feature of the Salicaceae SINEs is the contrast of the conservation of 5’ start motifs across species and SINE families compared to the high variability of
3’ ends within the SINE families, differing in sequence and length, presumably resulting from mutations in the poly(A) tail as a possible route for SINE elongation. Periods of increased transpositional activity promote the dissemination of novel 3’ ends. Thereby, evolutionarily older motifs are displaced leading to various 3’ end subpopulations within the SaliS families. Opposed to the PoaS families with a largely equal ratio of poly(A) to poly(T) tail SINEs, the SaliS families are exclusively terminated by adenine stretches.
Among retrotransposon-based markers, SINEs are highly suitable for the development of molecular markers due to their unidirectional insertion and random distribution mainly in euchromatic genome regions, together with an easy and fast detection of the heterogeneous SINE families. As a prerequisite for the development of SINE-derived inter-SINE amplified polymorphism (ISAP) markers, 13 novel Theaceae SINE families (TheaS-I - TheaS-VII, TheaS-VIII.1 and TheaS-VIII.2, TheaS-IX - TheaS-XIII) were identified in the angiosperm tree species Camellia japonica. Moreover, six Pinaceae SINE families (PinS-I.1 and PinS-I.2, PinS-II – PinS-VI) were detected in the gymnosperm species Larix decidua. Compared to the SaliS and PoaS families, structural relationships are less frequent within the TheaS families and absent in the PinS families.
The ISAP analysis revealed the genetic identity of Europe’s oldest historical camellia (C. japonica) trees indicating their vegetative propagation from the same ancestor specimen, which was probably the first living camellia on European ground introduced to England within the 18th century. Historical sources locate the native origin of this ancestral camellia specimen either in the Chinese province Yunnan or at the Japanese Gotō Islands. Comparative ISAPs showed no accordance to the Gotō camellia sample pool and appropriate Chinese reference samples were not available. However, the initial experiments demonstrated the potential of ISAP to resolve variations among natural populations.
The ISAP application on angiosperm trees also concerned fast growing Populus clones grown in short rotation coppice plantations for energy production. The species-specific P. tremula ISAP primers might also be applied for the discrimination of hybrid poplar clones involving P. tremuloides genome
portions, since SINEs of these two species are highly related. However, due to lineage-specific SINE evolution during speciation, cross-species applications are generally only successful to limited extent. The analysis of poplar hybrids composed of P. maximowiczii with either P. trichocarpa or P. nigra based on P. tremula ISAP primers showed a strongly reduced resolution.
In forestry, hybrid larch (e.g. Larix × eurolepis) genotypes have to be selected from the offspring of Japanese (Larix kaempferi) and European larch (Larix decidua) crosses, as they exhibit superior growth rates compared to the parental species. Initial ISAP-based examinations of European larch genotypes provided less polymorphic banding patterns, probably resulting from general high levels of synteny and collinearities reported for gymnosperm species. Hence, the ISAP was combined with the AFLP technique to the novel marker system inter-SINE-restriction site amplified polymorphism (ISRAP). The amplicons originating from genomic regions between SINEs and EcoRI cleavage sites were visualized with the sensitive capillary gel electrophoresis. The ISRAP assays, based on EcoRI adapter primers combined with two different SINE-derived primers, resulted in a sufficient number of polymorphic peaks to distinguish the L. decidua genotypes investigated. Compared to ISAPs, the ISRAP approach provides the required resolution to differentiate highly similar larch genotypes.
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Chutimanitsakun, Yada. "From the Oregon Wolfe Barley to fall-sown food barley : markers, maps, marker-assisted selection and quantitative trait loci." Thesis, 2011. http://hdl.handle.net/1957/26225.
Full textAbstract:
Understanding complex traits is a fundamental challenge in plant genetics and a prerequisite for molecular breeding. Tools for trait dissection are markers, maps, and quantitative trait locus (QTL) analysis. Marker-assisted selection (MAS) is an application that integrates these tools. In this thesis research, a new sequence-based marker was evaluated, maps were constructed and used, and QTLs were detected using two types of populations. Marker-assisted selection was used to develop a novel class of barley. Restriction-site Associated DNA (RAD), a sequence based-marker technology, allows for simultaneous high-density single nucleotide polymorphism (SNP) discovery and genotyping. We assessed the value of RAD markers for linkage map construction using the Oregon Wolfe Barley (OWB) mapping population. We compared a RAD-based map to a map generated using Illumina GoldenGate Assay (EST-based SNPs). The RAD markers generated a high quality map with complete genome coverage. We then used the RAD map to locate QTL for agronomic fitness traits. A paper describing this research was published (Chutimanitsakun et al., 2011). Marker-assisted selection was used to rapidly develop fall-sown barley germplasm for human food uses. The target traits were high grain β-glucan, vernalization sensitivity (VS) and low temperature tolerance (LTT). The target loci were WX and VRN-H2. Marker-assisted selection was effective in fixing target alleles at both loci and waxy starch led to increase in grain β-glucan. Unexpected segregation at VRN-H1 and VRN-H3, revealed by genome-wide association mapping (GW-AM), led to unanticipated phenotypic variation in VS and LTT. We found that GW-AM is an efficient and powerful method for identifying the genome coordinates of genes determining target traits. Precise information is obtained with perfect markers; additional research may be needed when multiple alleles are segregating at target loci and significant associations are with markers in linkage disequilibrium (LD) with the target loci. A paper describing this research will be submitted for publication.Graduation date: 2012