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Viswanathan, Aparna. "Phylogenetic analysis of sclerospora graminicola using internal transcribed spaced region-2." Texas A&M University, 2003. http://hdl.handle.net/1969/394.
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Förster, Frank [Verfasser], and Thomas [Akademischer Betreuer] Dandekar. "Making the most of phylogeny: Unique adaptations in tardigrades and 216374 internal transcribed spacer 2 structures / Frank Förster. Betreuer: Thomas Dandekar." Würzburg : Universitätsbibliothek der Universität Würzburg, 2012. http://d-nb.info/1022061232/34.
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Tozkar, Ozge Cansu. "Comparative Sequence Analysis Of The Internal Transcribed Spacer 2 Region Of Turkish Red Pine (pinus Brutia Ten.) And Natural Aleppo Pine (pinus Halepensis Mill.) Populations From Turkey." Master's thesis, METU, 2007. http://etd.lib.metu.edu.tr/upload/2/12608313/index.pdf.
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ABSTRACT
COMPARATIVE SEQUENCE ANALYSIS OF THE INTERNAL TRANSCRIBED SPACER 2 REGION OF TURKISH RED PINE (Pinus brutia TEN.) AND NATURAL ALEPPO PINE (Pinus halepensis MILL.) POPULATIONS FROM TURKEY
Tozkar, Özge M.S., Department of Biology Supervisor: Prof. Dr. Zeki Kaya April, 2007, 107 pages Turkish red pine (Pinus brutia) is wide-spread and an important forest tree species in Turkey, occurring mainly in southern, western and north-western Turkey and as small isolated populations in the Black Sea region. Aleppo pine (Pinus halepensis) has naturally found only in Adana and Mugla provinces as small population in mixture with Turkish red pine. Although Turkish red pine and Aleppo pine are morphologically different, Turkish red pine has been regarded as subspecies of Aleppo pine by some taxonomists due to occurrence of natural hybridization between these two species. However, the phylogenic relationship between these species needs to be explored further. In the present study, by sampling overlapped populations of both species from Mugla and Adana provinces (4 populations of Turkish red pine and 3 populations of Aleppo pine), internal transcribed spacer (ITS) region of ribosomal DNA were comparatively studied with sequence analysis. Although ITS1, 5.8s and ITS2 regions of ribosomal DNA were studied with ITS primers, only ITS2 region was successfully amplified with polymerase chain reaction (PCR). The complete data set for this region was analysed using MEGA3.1 and Arlequin softwares. Analysis of molecular variance (AMOVA) demonstrated the highest genetic differentiation between Turkish red pine and Aleppo pine in Mugla with 100 percentage of variation. AMOVA analysis also indicated the possibility of low-level migration of genes between Turkish red pine and Aleppo pine populations in Adana with 50.65 percent of molecular variance. Haplotype comparison revealed that two major haplotypes were represented Based on the results of ITS2 region sequence analysis, Turkish populations of Aleppo pine and Turkish red pine populations could not be fully differentiated. In Mugla province Turkish red pine and Aleppo pine revealed more differentiation due to reproductive isolation. But in Adana province, two species shared more common genetic background due to possible hybridization. Since ITS2 region of nuclear ribosomal DNA revealed a few variable and parsimony informative sites for both species, thus, only ITS2 region of ribosomal DNA does not appear to be sufficient for fully resolving genetic relationships between Turkish red pine and Aleppo pine populations. Further studies including ITS1 and 5.8s regions of ribosomal DNA and populations included from major Aleppo pine distribution areas will be useful to understand the evolutionary relationship between Aleppo pine and Turkish red pine populations in Turkey.
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Cheung, Mei, and 張微. "Internal transcribed spacer as the DNA barcode for pathogenic fungi." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2014. http://hdl.handle.net/10722/206495.
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Identification of pathogenic fungi isolated from clinical specimens in clinical microbiology laboratories is primarily based on observing fungal phenotypic structures under the microscope and performing biochemical tests for fungal cultures. This conventional method is very time-consuming and labor-dependent. It usually requires several weeks for the fungi to grow sufficiently on culture media, and the identification processes on fungal phenotypic structure rely very much on experienced staff. Therefore, a more accurate and rapid method for pathogenic fungal identification is necessary for clinical laboratories to get abreast of modern development.
Gene sequencing and phylogenetic analysis targeting the internal transcribed spacer (ITS) region in the fungal genomes are the most commonly used molecular methods for fungal identification. Because of the optimal inter and intra-species variation property of the ITS region, it can act as the DNA barcode to identify fungi to the species level. In this study, 33 clinical fungal isolates were identified by both phenotypic method and ITS sequencing. The results showed that 23 isolates were successfully identified to thespecies level by both phenotypic and molecular methods. Moreover, five isolates were only identified to the genus level by phenotypic method, but they could be successfully identified to the species level by ITS sequencing. However, five isolates have not been differentiated because there were mismatched results from phenotypic and sequencing methods. It may be due to the limitation of sequencing method on some fungal species. Building up a more comprehensive database or setting up a standard platform to guide the molecular process may help improve the performance of molecular method.
To conclude, molecular method is a rapid and reliable way for fungal identification because ITS region acts as the DNA barcode for pathogenic fungi.published_or_final_version
Medical Sciences
Master
Master of Medical Sciences
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Bodo, Slotta Tracey A. "Phylogenetic Analysis of Iliamna (Malvaceae) Using the Internal Transcribed Spacer Region." Thesis, Virginia Tech, 2000. http://hdl.handle.net/10919/33211.
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The genus Iliamna Greene has a taxonomically complex history. Since its desciption in 1906, the genus was not recognized for some time, several species were initially placed into other genera, and the species status of a few was questioned. Today, eight species of Iliamna are recognized. Six species are located in western North America and two are found isolated to the east. Species in Iliamna are very similar morphologically with only a few characters distinguishing several as separate entities. The need for systematic study became apparent since all but one species are considered rare or endangered. Also, the differentiation between two endangered species, I. corei and I. remota, was unclear in a previous study using random amplified polymorphic DNA fragments. Of the western species, four overlap in distribution (I. crandallii, I grandi ora, I. longisepala, and I. rivularis) and their recognition as separate species has been questioned. The focus of this study was to develop a phylogeny for Iliamna using sequences from the internal transcribed spacer region (ITS) of the nuclear ribosomal
RNA genes in order to determine its biogeographical and evolutionary history. Cladistic analysis was performed and the resulting phylogeny is presented. The ITS data provide new insights in the origination of the genus and its distribution. In Iliamna, the ITS region is 677 base pairs long with 120 sites providing information in the formation of phylogenetic trees. Iliamna forms a well-supported clade distinct from related genera and is monophyletic. Three well-supported groups are formed. One contains representatives from the Pacific Northwest. Another contains all of the remaining species with the third clade nested therein. This last clade contains the two eastern species, I. corei and I. remota, but there is not enough variability to support the divergence of these taxa as distinct species. There is also not sufficient variability in the ITS region to identify the western species I. crandallii, I. grandi ora, I. longisepala and I. rivularis as distinct entities.Master of Science
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Hao, Gang, and 郝剛. "Molecular phylogeny of the illiciales based on internal transcribed spacer sequences of ribosomal DNA." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 1999. http://hub.hku.hk/bib/B31238567.
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Hao, Gang. "Molecular phylogeny of the illiciales based on internal transcribed spacer sequences of ribosomal DNA /." Hong Kong : University of Hong Kong, 1999. http://sunzi.lib.hku.hk/hkuto/record.jsp?B21029027.
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Ragozine, Vincent. "Analyses of ribosomal DNA internal transcribed spacer sequences from : Juglans nigra and leaf-associated fungi in Zoar Valley, NY /." Connect to resource online, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=ysu1211685563.
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Ragozine, Vincent Kyle. "Analyses of ribosomal DNA internal transcribed spacer sequences from Juglans nigra and leaf-associated fungi in Zoar Valley, NY." Youngstown State University / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=ysu1211685563.
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Heidemann, Stefan [Verfasser]. "DNA-Polymorphismen der internal transcribed spacer-Region zur Differenzierung anthropophiler und zoophiler Spezies von Trichophyton mentagrophytes sensu lato / Stefan Heidemann." Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2010. http://d-nb.info/1024334767/34.
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Nyarku, Rejoice E. "Development and analytical validation of a genus-specific Brucella real-time PCR assay targeting the 16S-23S rDNA internal transcribed spacer." Diss., University of Pretoria, 2020. http://hdl.handle.net/2263/77428.
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Brucellosis is an economically important bacterial disease of both animals and humans. In sub-Saharan Africa, the diagnosis of the disease remains a challenge. Brucellosis is underreported in South Africa, due to inconsistency in reports of bacteriological and serological tests, which lack adequate sensitivity and specificity in the diagnosis of the disease. They also are ineffective in confirming brucellosis during early stages of the disease.
The aim of this study was to develop a 16S-23S ribosomal deoxyribonucleic acid (rDNA) internal transcribed spacer (ITS) quantitative polymerase chain reaction (qPCR) assay for early diagnosis of brucellosis and as a rapid screening tool. To achieve this, blood, milk and tissue samples were spiked with B. abortus biovar (bv.) 1 (B01988-18 strain) to determine the analytical sensitivity and specificity of the assay. The efficiency was 105% in tissue, 99% in blood, and 93% in milk. The 95% limit of detection (LOD) of the ITS qPCR assay was highest in tissue, followed by blood, then milk; thus (1.45, 13.30 and 45.54 bacterial genome copies/PCR reaction).
Furthermore, the diagnostic performance of the assay was compared to the Brucella cell surface protein real time polymerase chain reaction (BCSP31 qPCR) assay. Out of 56 aborted foetal tissue samples from bovine, ovine and caprine, 33% (19/56) were positive for Brucella spp. The sensitivity and specificity of the ITS qPCR assay were 87% and 95% respectively, compared to the 92% and 89% for the BCSP31 qPCR assay and 47% and 55% for bacterial culture, respectively. The ITS qPCR gave earlier CT’s with a difference in CT (ΔCT) between ITS and BCSP31 ranging between 7.1 and 3.24.
The assay was efficient, sensitive and specific. It detected as little as 1.45 bacterial genome copies/PCR reaction in tissue, making this assay a valuable tool in early detection of the presence of the Brucella pathogen. It is sensitive and specific in the diagnosis of brucellosis.Dissertation (MSc)--University of Pretoria, 2019.
Veterinary Tropical Diseases
MSc
Unrestricted
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Ross, Sara J. "Molecular Phylogeography and Species Discrimination of Freshwater Cladophora (Cladophorales, Chlorophyta) in North America." Thesis, University of Waterloo, 2006. http://hdl.handle.net/10012/2977.
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Cladophora is a widespread freshwater filamentous cholorophyte genus and is frequently observed in eutrophic waters where it can produce large nuisance blooms. These blooms can have direct impacts on water intake for power generation, irrigation canals and can be aesthetically unpleasant. Much of the ecological and physiological studies on Cladophora have assumed that the populations of this genus in North America belong to the species Cladophora glomerata. However, this has never been tested despite that it is well documented that identifying freshwater Cladophora to the species level is difficult due morphological variability under different ecological conditions. In addition, the species epithets for freshwater Cladophora are based on European collections and it is not clear if these should be applied to North America. This study examines approximately 40 collections of Cladophora from the Laurentian Great Lakes and 43 from various locations in North America ranging from the Northwest Territories to Puerto Rico. Initially we determined the nucleotide sequences of the internal transcribed spacer (ITS) region of the nuclear ribosomal cistron and observed sequence divergence to be low (0-3%), demonstrating an inability for this marker to resolve species delineation as divergence of this region was low. Amplification of the inter-simple sequence repeat (ISSR) regions were used to analyze microsatellite motif frequency throughout the genome to evaluate the biogeography relationships, including diversity, of freshwater Cladophora sp. five different primers were used on 70 individuals. UPGMA analyses of the presence/absence of bands demonstrate that each of the Great Lake populations separate into groups according to the Lake they were initially sampled from. However, collections from North America are highly variable and do not form well supported biogeographic clades. In addition, these collections appear to be distinct from type cultures of freshwater Cladophora from Europe. Supplementary morphological analysis using suggested taxonomically valid criterion (length and diameter of main axis, ultimate branch, and apical cell) none were able to differentiate Great Lake populations.
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Kitz, Leilani. "Evaluation of Downy Mildew (Peronospora farinosa f.sp. chenopodii) Resistance among Quinoa Genotypes and Investigation of P. farinosa Growth using Scanning Electron Microscopy." Diss., CLICK HERE for online access, 2008. http://contentdm.lib.byu.edu/ETD/image/etd2537.pdf.
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Sidiq, Farida P. "Identification of culture-negative fungi in blood and respiratory samples." Bowling Green State University / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1393517514.
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Meireles, Jose Eduardo de Carvalho. "Revisão taxonomica e filogenia de Poecilanthe s.l. (Leguminosae, Papilionoideae, Brongniartieae)." [s.n.], 2007. http://repositorio.unicamp.br/jspui/handle/REPOSIP/314808.
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Orientador: Ana Maria Goulart de Azevedo TozziDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Poecilanthe (Leguminosae, Papilionoideae, Brongniartieae) é em gênero sul-americano que inclui atualmente dez espécies. A heterogeneidade morfológica e química encontrada em Poecilanthe dificulta sua circunscrição e coloca em dúvida sua monofilia. Além disso, limites interespecíficos imprecisos e falta de chave de identificação dificultam o reconhecimento das espécies. Este trabalho tem como objetivos testar a monofilia de Poecilanthe e estabelecer as relações entre suas espécies, bem como revisar a taxonomia do gênero. Para tanto, uma análise filogenética de máxima parcimônia baseada em caracteres morfológicos e seqüências de ITS/5.8S (nrDNA) foi realizada. Como subsídio para a análise cladística, foi feito um estudo sobre a morfologia das sementes e embriões de Poecilanthe, que resultou no reconhecimento de quatro padrões distintos de morfologia. Os resultados da filogenia mostram que Poecilanthe não é um gênero monofilético, sendo composto por três clados parafiléticos em relação à tribo. Estes três clados foram caracterizados morfologicamente e considerados como gêneros distintos. Poecilanthe é recircunscrito para incluir apenas as espécies extra-amazônicas (Poecilanthe s.s.), compreendendo então seis espécies. O gênero Amphiodon é restabelecido, e P. ovalifolia combinada neste. Um gênero novo é descrito para incluir P. amazonica e P. hostmannii. Cada um destes gêneros foi tratado taxonomicamente, constando em cada tratamento descrições, ilustrações e chave para a identificação das espécies
Abstract: The genus Poecilanthe (Leguminosae, Papilionoideae, Brongniartieae) currently comprises ten South-American species. The morphological and chemical diversity that is found within this genus renders its circumscription imprecise and brings Poecilanthe¿s monophyly into question. This work aims to test the monophyly of Poecilanthe and to revise the taxonomy of the genus. A parsimony analysis based on both morphological and ITS/5.8S data was carried out. In order to provide characters to the cladistic analysis, the morphology of the seeds and embryos of Poecilanthe was analyzed, and resulted in the identification of four different morphological patterns. The phylogeny does not support Poecilanthe as monophyletic, but resolves three different well-supported lineages that are paraphyletic with respect to the tribe. These clades are morphologically characterized and ranked at the generic level. Poecilanthe is recircumscribed to include the six extra-Amazonian species only. The genus Amphiodon is reinstated and P. ovalifolia is combined. Poecilante amazonica and P. hostmannii are segregated into a new genus. Each genus was revised and descriptions, illustrations and identification key for the species are presented.
Mestrado
Biologia Vegetal
Mestre em Biologia Vegetal
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Turenne, Christine Yvette. "The use of the 16S rRNA gene and the internal transcribed spacer, ITS2, region for rapid and specific identification of bacteria and fungi from clinical specimens." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp03/MQ32970.pdf.
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Vargas, Mariana L. "Host-parasite coevolution in New Zealand: how has Odontacarus, a mite with a free-living stage in its life-cycle, coevolved with its skink host?" Lincoln University, 2006. http://hdl.handle.net/10182/1072.
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The effect of a free-living stage in host-parasite coevolution: a skink mite phylogenetic study in New Zealand. During the last decade, phylogenetic trees have even been used to compare ecologically related taxa such as parasites and their hosts, and are used to determine their level of coevolution or reciprocal adaptation in time. Diverse coevolutionary events have been detected for this ecological association, where generally the parasite has been regarded as one that feeds exclusively on the host and is likely to cospeciate with it. A different coevolutionary pattern might occur when the parasite has a free-living stage in its life cycle, in which the parasite may have the opportunity to abandon its host and successfully colonise a new species (host-switching) making cospeciation less likely. Many New Zealand skinks are infested with a parasitic mite, Odontacarus sp. (Prostigmata: Leeuwenhoekiidae), which becomes free-living as an adult. The genetic variation of these mites found on four hosts was analyzed for host- parasite coevolutionary events. The hosts were the McCann’s skink and the common skink in coastal Birdling Flat, Canterbury, plus these species and the Grand and Otago skinks in Macraes Flat, Central Otago, South Island, New Zealand. The genetic variation of fast evolving nuclear Internal Transcribed Spacers 2 and mitochondrial Cytochrome c Oxidase I in Odontacarus mites found on these hosts was determined by PCR and DNA sequencing and phylogenetic trees were built using the computer programs PAUP*4 and MrBayes 3. The results show that mite haplotypes only had a significant geographical division and no host-related differences. In Birdling Flat, the COI haplotypes were represented in two groups that infested both regional hosts and had 5.7 % divergence. The same individual mites belonged to a single ITS 2 haplotype, thus indicating a historical geographical division between two populations that now interbreed successfully. The Macraes Flat mites were divided into two COI haplotypes with 2.4% divergence and internal nodes, which showed greater genetic variability than the Birdling Flat populations. The Macraes Flat mites formed two ITS 2 haplotypes with 6% divergence. This greater geographical structure of the Otago mites is probably due to the older age of the mainland area compared to the recently exposed coastal locality of Birdling Flat. The COI haplotypes from the two different regions had a mean distance of 15.5%, with an earlier divergence time than that known for the hosts. For both genes, the haplotypes from different regions had 100% bootstrap support and the parasite showed no host specificity. Mites of the different COI and ITS haplotypes were found on most of the host species that were sampled in Canterbury and Otago. The results of this study suggest that a free-living stage in a parasite’s life cycle can favour coevolutionary events such as inertia (failure to speciate) and host-switching, probably as a result of resource-tracking of the parasite. NB: Electronic files contained on CD to accompany print copy are not included with this version of the thesis.
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Magalon, Hélène. "Dispersion du corail Pocillopora meandrina et de ses algues symbiotiques en Polynésie Française." Paris 6, 2005. http://www.theses.fr/2005PA066100.
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Woods, Kristi Yvonne. "Nymphaea odorata (Water-lily, Nymphaeaceae): Analyses of molecular and morphological studies." Thesis, Virginia Tech, 2003. http://hdl.handle.net/10919/41234.
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Molecular and morphologic studies were used to determine the evolution, classification and differentiation of Nymphaea odorata. Molecular analyses of the nuclear internal transcribed spacer (ITS) region, the chloroplast trnL-F region, and inter-simple sequence repeat (ISSR) markers determined the variation present between and within two species of Nymphaea. The ITS region resulted in a phylogeny depicting strong separation between species (N. mexicana and N. odorata) and some separation between N. odorataâ s subspecies. The ITS region contained polymorphisms, which upon SAHN clustering and principle coordinate (PCOA) and minimum spanning tree (MST) analyses produced groups similar to the clades in the ITS phylogeny. Sixteen accessions were chosen for trnL-F analysis, where a subspecies-specific molecular marker was found. In most accessions the marker confirmed the original subspecies classification. Molecular analyses using ISSRs characterized among population variation in N. odorata and N. mexicana using five primers. ISSR markers among populations were highly variable within a species and were used in UPGMA, PCOA and MST analysis, which resulted in separation between the subspecies.
Both univariate and multivariate analyses were performed on quantitative and qualitative morphological characters. An analysis of variance resulted in six morphological characteristics that were statistically significant (P< 0.05), the majority being leaf blade characteristics. Multivariate statistics of principle component analysis and discriminate analysis resulted in groups for each subspecies, both emphasized the importance of quantitative leaf blade characteristics. Overall, both morphology and molecular characteristics supported the classification of subspecies for ssp. odorata and ssp. tuberosa, due a lack of strong segregation of characteristics.Master of Science
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Müller, Juceli. "Qualidade fisiológica e associação de Fusarium spp. a sementes de sorgo sacarino." Universidade Federal de Santa Maria, 2017. http://repositorio.ufsm.br/handle/1/11796.
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The present work aims to determine the physiological and sanitary quality of Sweet sorghum (Sorghum bicolor (L.) Moench) seeds, as well as to identify pathogens associated with seed, their transmission to seedlings and the subsequent pathogenicity of isolates obtained, In addition, molecularly identify the fungal species pathogenic to this crop. The experiments were carried out in the Teaching and Seed Research Laboratory (TSRL), of the Plant Engineering Department; In the Elocy Minussi Phytopathology Laboratory, Department of Plant Protection, and at the Biological Institute of São Paulo. Sweet sorghum seeds were used, all without chemical treatment. Sanitary quality was evaluated by sanity test, and physiological characteristics by germination and vigor tests (seedling length, dry mass, emergence, rate of emergence and accelerated aging). It was performed the test of transmission of the pathogens associated to the seeds and the subsequent pathogenicity of the obtained isolates, culminating with the molecular characterization of the identified pathogens, in which were sequenced the Internal Transcribed Spacer (ITS) genomic regions and the Elongation Factor 1 - alpha (TEF1-α) gene. The design used was the completely randomized design, with four cultivars of Sweet sorghum (BRS 506, F19, BRS 511 and BRS 509); For the evaluation of pathogenicity, the factorial scheme is represented by four cultivars and three isolates of Fusarium spp., besides the witness. The seeds of the BRS 509 cultivar were considered to have lower physiological quality than the other cultivars. The DNA sequencing allowed identifying the Fusarium thapsinum species as a pathogenic agent in the sweet sorghum crop, and proven its transmission via seeds.O presente trabalho teve como objetivo determinar a qualidade fisiológica e sanitária de sementes de cultivares de sorgo sacarino (Sorghum bicolor (L.) Moench), bem como identificar os patógenos associados à semente, sua transmissão às plântulas e a posterior patogenicidade de isolados obtidos, além disso, identificar molecularmente as espécies fúngicas patogênicas a esta cultura. Os experimentos foram realizados no Laboratório Didático e de Pesquisas em Sementes (LDPS), do Departamento de Fitotecnia; no Laboratório de Fitopatologia Elocy Minussi, do Departamento de Defesa Fitossanitária e, no Instituto Biológico de São Paulo. Foram utilizadas sementes de sorgo sacarino, todas sem tratamento químico. A qualidade sanitária foi avaliada pelo teste de sanidade, e as características fisiológicas por meio dos testes de germinação e de vigor (comprimento de plântulas, massa seca, emergência, índice de velocidade de emergência e envelhecimento acelerado). Foi realizado o teste de transmissão dos patógenos associados à semente e a posterior patogenicidade dos isolados fúngicos obtidos, culminando com a caracterização molecular dos patógenos identificados, na qual foram sequenciadas as regiões genômicas Internal Transcribed Spacer (ITS) e o gene do fator de elongação 1-α (TEF1α). O delineamento utilizado foi o inteiramente casualizado com quatro cultivares de sorgo sacarino (BRS 506, Fepagro 19, BRS 511 e BRS 509); já para a avaliação da patogenicidade, o esquema fatorial foi representado pelas quatro cultivares e três isolados de Fusarium sp., além da testemunha. As sementes da cultivar BRS 509 foram consideradas de qualidade fisiológica inferior as demais cultivares. O sequenciamento de DNA permitiu identificar a espécie Fusarium thapsinum como agente patogênico na cultura do sorgo sacarino, sendo comprovada sua transmissão via sementes.
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Glass, Pamela Michele. "Evidence of Ecological Speciation in Phacelia." Digital Commons @ East Tennessee State University, 2007. https://dc.etsu.edu/etd/2143.
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Phacelia purshii Buckley and P. fimbriata Micheaux are two species that are nearly morphologically indistinguishable. Seed germination experiments showed that the high elevation endemic, P. fimbriata requires lower temperatures to trigger germination. Following interspecific crosses, pollen tubes enter ovules and maternal tissue of the gynoecium matures but hybrid diploid and triploid organs fail to develop. DNA sequences from the ribosomal DNA internal transcribed region showed that P. fimbriata and P. purshii comprise a monophyletic clade but that P. fimbriata is more differentiated from related species. In contrast, P. purshii supported significantly higher levels of intraspecific polymorphism. Phacelia fimbriata and P. purshii are sister species with similar morphology but they are unable to hybridize, they are differentiated in physiological characteristics related to environment, and they inhabit different elevations. This pattern of relationship and differentiation suggests P. fimbriata may be the product of ecological speciation.
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Marques, Cálita Pollyanna. "Padronização da análise dos produtos da PCR/RFLP que amplifica os genes Ribossomal Internal transcribed spacer (ITS) e Glucose-6- phosphate dehydrogenase (G6PD) para identificação de Leishmania spp. em gel de poliacrilamida." Universidade Federal de Goiás, 2017. http://repositorio.bc.ufg.br/tede/handle/tede/7789.
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American Cutaneous Leishmaniasis (LTA) is caused by protozoa of the genus Leishmania and it attacks the skin and/or mucous membranes. It is an endemic zoonosis in Brazil being notified in all the Regions. There is a relation between the species of parasite and the clinical manifestations, contributing to the diagnostic complexity, which up to the present, did not exist species-specific diagnosis. In the present work, the proposal was to standardize the analysis of the PCR/RFLP products to the ITS and G6PD genes in polyacrylamide gel. In order to standardize PCR/RFLP analysis on the ITS and G6PD genes, DNA from the reference Leishmania spp. Sample was used and 5%, 6%, 7% 8% and 10% polyacrylamide were tested. The protocol design considered the size of the amplicons generated by the PCR / RFLP, correlating the concentration of the polyacrylamide. To standardize the electrophoresis time the migration of the dyes used in the DNA sample buffer was monitored. To validate the technique, we selected 521 samples from Tocantins patients fixed on slides and 43 samples of parasite isolates stored in Leishbank. The samples from the patients from Tocantins were analyzed by PCR/RFLP to the kDNA and were selected to those positive for the validation. The concentration of the matrix for analysis of the PCR and RFLP products to the ITS gene was standardized in 6% and 8%, respectively, and distinguished the species L. (L.) amazonensis, but the species L. (V.) braziliensis, L. (V.) guyanensis and L. (V.) lainsoni were not distinguished among themselves, occurring the same between L. (V.) naiffi and L. (V.) shawi. The concentration of the matrix for analysis of the PCR products to the G6PD gene was standardized in 6% and distinguished L. (V.) braziliensis from the others. The validation of the polyacrylamide gel analysis of the PCR products to the ITS target from samples of patients fixed in slides was not performed, since of the 256 samples analyzed only 4 amplified. The validation of the analysis of the products of the PCR to the G6PD target in polyacrylamide gel from 48 samples fixed in slides characterized 81% as L. (V.) braziliensis and, characterized 63% of the isolates of parasites as L. (V.) braziliensis. The PCR/RFLP analyzes performed to discriminate Leishmania spp. are complementary and thus contribute to the species-specific diagnosis of LTA.
A Leishmaniose Tegumentar Americana (LTA) é causada por protozoários do gênero Leishmania e acomete pele e/ou mucosas. É uma zoonose endêmica no Brasil sendo notificada em todas as Regiões. Há uma relação entre a espécie de parasito e as manifestações clínicas, contribuindo à complexidade diagnóstica, que até o presente, não existi o diagnóstico espécie-específico. No presente trabalho, a proposta foi padronizar a análise dos produtos da PCR/RFLP aos genes ITS e G6PD em gel de poliacrilamida. Para padronizar as análises dos produtos da PCR/RFLP aos genes ITS e G6PD, foram utilizadas DNA da amostra de Leishmania spp., de referência, sendo testada a poliacrilamida na concentração de 5%, 6%, 7% 8% e 10%. O delineamento do protocolo considerou o tamanho dos amplicons gerados pela PCR/RFLP, correlacionando-o a concentração da poliacrilamida. Para padronizar o tempo da eletroforese foi monitorada a migração dos corantes usados no tampão da amostra de DNA. Para validar a técnica, foram selecionadas 521 amostras de pacientes tocantinenses fixadas em lâminas e 43 amostras de isolados de parasitos armazenados no Leishbank. As amostras dos pacientes tocantinenses foram analisadas pela PCR/RFLP ao kDNA e foram selecionadas àquelas positivas para a validação. A concentração da matriz para analise dos produtos da PCR e RFLP ao gene ITS ficou padronizada respectivamente em 6% e 8% e distinguiu a espécie L.(L.) amazonensis, porém as espécies L. (V.) braziliensis, L. (V.) guyanensis e L. (V.) lainsoni não foram distinguidas entre si, ocorrendo o mesmo entre L. (V.) naiffi e L. (V.) shawi. A concentração da matriz para analise dos produtos da PCR ao gene G6PD ficou padronizada em 6% e distinguiu L. (V.) braziliensis das demais. A validação da análise em gel de poliacrilamida dos produtos da PCR ao alvo ITS a partir de amostras de pacientes fixadas em lâminas não ser realizada, pois das 256 amostras analisadas somente 4 amplificaram. A validação da análise dos produtos da PCR ao alvo G6PD em gel de poliacrilamida a partir de 48 amostras fixadas em lâminas caracterizou 81% como L. (V.) braziliensis e, caracterizou 63% dos isolados de parasitos como L. (V.) braziliensis. As análises pela PCR/RFLP realizadas para discriminar Leishmania spp. se complementam e, assim, contribuem para o diagnóstico espécie-específico da LTA.
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Marrelli, Mauro Toledo. "Anopheles oswaldoi (Diptera, Culicidae): análise do segundo espaçador interno transcrito (ITS2) do DNA ribossômico e da susceptibilidade à infecção com Plasmodium vivax." Universidade de São Paulo, 2000. http://www.teses.usp.br/teses/disponiveis/42/42135/tde-30032001-141738/.
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Resultados anteriores sugerem que existem diferenças biológicas entre espécimens de Anopheles oswaldoi capturados no Estado do Acre e os do Estado de Rondônia, Brasil. Esta espécie tem sido apontada como um importante vetor de malária em localidades do Peru e Acre. Entretanto, em Rondônia, somente um número pequeno de A. oswaldoi alimentados em pacientes com malária desenvolveram infecção nas glândulas salivares. Além disso, há suspeita de que espécimens identificados como A. oswaldoi, capturados em áreas abertas em Costa Marques, Rondônia, são na verdade A. konderi, e que A. oswaldoi sensu stricto estaria restrito a áreas de florestas. Estes dados, juntamente com as dificuldades encontradas na identificação de anofelinos do grupo Oswaldoi, baseadas em critérios morfológicos, sugerem que espécimens de A. oswaldoi são membros de um complexo de espécies crípticas. A distinção de espécies crípticas de insetos vetores é de grande importância, já que diferentes membros em um complexo podem exibir diferenças na ecologia, capacidade vetorial e resposta a medidas de controle. Análise de sequências de DNA, particularmente da região do ITS2 do cistron do DNA ribossômico, tem sido usada como um caracter diagnóstico em alguns grupos de espécies crípticas, tornando-se um instrumento para estudos taxonômicos e filogenéticos. A primeira parte deste estudo teve o objetivo de determinar as diferenças encontradas nas sequências de ITS2 de espécimens de A. oswaldoi capturados em várias localidades da América do Sul. As regiões dos ITS2 destes anofelinos foram amplificadas usando oligonucleotídeos iniciadores conservados das regiões 5.8S e 28S e os produtos de PCR foram clonados e sequenciados. Os ITS2 de todos os mosquitos capturados tiveram tamanho aproximado de 350 nucleotídeos, com aproximadamente 53% de conteúdo de GC. Análise do alinhamento destas sequências, mostrou similaridade variando entre 87% e 100%, e a análise de uma árvore de similaridade, neighbor-joining, produzida com p-distance usando as diferenças nas sequências de ITS2, separou estes espécimens em quatro grupos. Um deles está provavelmente relacionado ao A. oswaldoi sensu stricto, e um outro pode estar relacionado à espécie A. konderi. Os outros dois grupos podem corresponder a espécies cuja identificação morfológica permanece para ser esclarecida no complexo A. oswaldoi. Estes dados são evidências de que espécimens de A. oswaldoi estão incluídos em um complexo de espécies crípticas e que identificação por DNA pode resolver estas questões taxonômicas. A. konderi tem sido considerado uma sinonímia de A. oswaldoi. Embora estágios adultos e imaturos destas espécies de anofelinos apresentem características morfológicas idênticas, aspectos encontrados na genitália masculina podem distinguir estes taxa. A segunda parte deste estudo foi conduzida com o objetivo de comparar a susceptibilidade de A. oswaldoi s.s. e de A. konderi à infecção com Plasmodium vivax. A susceptibilidade foi baseada na proporção de mosquitos com oocistos e esporozoítos. Os anofelinos foram capturados no Acre e em Rondônia para obtenção de progênies F1. Após a emergência dos adultos, as genitálias masculinas dos mosquitos de cada progênie foram dissecadas e examinadas. Todas progênies originadas dos mosquitos capturados em Rondônia corresponderam a A. konderi, enquanto que cerca de 85,0% das progênies do Acre eram A. oswaldoi s.s.. Estas progênies F1 de A. oswaldoi s.s., A. konderi e A. darlingi foram alimentadas simultaneamente com sangue infectado com P. vivax. Estes mosquitos foram dissecados 10-12 dias após infecção e examinados para verificação da presença de oocistos e esporozoítos. Tanto A. oswaldoi s.s. como A. konderi apresentaram oocistos nos tratos digestivos, entretanto, a porcentagem de tratos digestivos positivos para oocistos foi maior em A. oswaldoi s.s. (13,8%) do que em A. konderi (3,3%). Esporozoítos foram encontrados somente nas glândulas salivares de A. oswaldoi s.s., com 6,9% de positividade. As taxas de infecção nos controles A. darlingi foram de 22,5% a 30,0%, para ambos oocistos e esporozoítos. Estes resultados indicam que A. oswaldoi s.s. pode transmitir P. vivax e sugerem que esta espécie é mais susceptível que A. konderi. Embora A. oswaldoi s.s. seja uma espécie exofílica e zoofílica, este anofelino pode estar envolvido na transmissão da malária humana como parece estar ocorrendo no Estado do Acre.Previous results have suggested that biological differences could exist between specimens of Anopheles oswaldoi captured in the State of Acre and those from the State of Rondônia, Brazil. This species has been appointed as an important malaria vector in localities of Peru and Acre. However, in Rondônia, only a very low percentage of A. oswaldoi fed on malaria patients developed salivary gland infections. In addition, it was suspected that specimens identified as A. oswaldoi captured in open clearings in Costa Marques, Rondônia, were actually A. konderi, and that A. oswaldoi sensu stricto would be restricted to forested areas. These data, together with the difficulties found to identify anophelines of the Oswaldoi group based on morphologic criteria suggest that specimens of A. oswaldoi are members of a complex of cryptic species. The distinction of cryptic species of insects is of critical importance, since different members in a Complex could exhibit differences in ecology, vectorial capacity and response to control measures. DNA sequence analysis and particularly that of the ITS2 region of the rDNA cistron has provided diagnostic characters in some groups of cryptic species becoming a general tool for taxonomic and phylogenetic studies. The first part of the present study was undertaken to determine the extent of differences over the ITS2 sequence of specimens of A. oswaldoi s.l. captured in several localities of South America. The ITS2 of these anophelines were amplified using conserved primers of the 5.8S and 28S regions, cloned and sequenced. The lengths of ITS2 of all mosquitoes captured were about 350 nucleotides, with about 53% GC content. Analysis of the alignment of the sequences, which showed that the similarity varied between 87% and 100%, and analysis of a neighbor-joining tree produced with p-distance using the ITS2 sequences, separated these specimens in four groups. One of them is probably related to A. oswaldoi sensu stricto, and another one can possibly be related to A. konderi. The other two groups may correspond to species the morphologycal identification of which remains to be clarified in the A. oswaldoi complex. These data are evidences that specimens of A. oswaldoi are included in a complex of cryptic species and that the DNA identification could solve some taxonomic questions. A. konderi has been currently considered to be a synonym of A. oswaldoi. Although adults and immature stages of both these anopheline species have identical morphological characters, features of the male genitalia can distinguish these taxa. The second part of this study was conducted in order to compare the susceptibility of A. oswaldoi s.s. and of A. konderi to infection by Plasmodium vivax. The susceptibility was based on the proportion of mosquitoes with oocysts and sporozoites. Anophelines were captured in the State of Acre and Rondônia and used to obtain F1 progenies. After emergency of adults, male genitalia of mosquitoes of each family were dissected. All families originated from mosquitoes captured in Rondônia corresponded to A. konderi, while about 85.0% of the families from Acre were A. oswaldoi s.s.. F1 progeny of field-captured A. oswaldoi s.s., A. konderi and A. darlingi were fed simultaneously on P. vivax infected blood. Mosquitoes were dissected on day 10-12 after infection and examined for the presence of oocysts and sporozoites. Both A. oswaldoi s.s. and A. konderi developed oocysts in midguts, however, the percentage of oocyst-positives in A. oswaldoi s.s. (13.8%) was higher than in A. konderi (3.3%), and only A. oswaldoi s.s. developed salivary infection with sporozoites (6.9% of positivity). Infection rates in A. darlingi ranged from 22.5% to 30.0% for both oocysts and sporozoites. These results indicate that A. oswaldoi s.s. can transmit P. vivax and suggest that it is more susceptible than A. konderi. Although A. oswaldoi s.s. is an exophilic and zoophilic species, it may be involved in human malaria transmission as it seems to be occuring in the State of Acre.
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Popp, Magnus. "Disentangling the Reticulate History of Polyploids in Silene (Caryophyllaceae)." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis : Univ.-bibl. [distributör], 2004. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-3918.
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González, Gaarslev Natalia. "Growth and biodegradation by Sporidiobolales yeasts in vanillin-supplemented medium." Thesis, Högskolan i Gävle, Avdelningen för arbets- och folkhälsovetenskap, 2017. http://urn.kb.se/resolve?urn=urn:nbn:se:hig:diva-24678.
Full textAbstract:
Studies of biodegradation in lignins by basidiomycetes yeasts show the conversion of lignin in various degradation products among which vanillin, a valuable substance, suggested to be a strong inhibitor of both fermentation and growth of yeasts, stands. Sporidiobolales yeasts used in these experiments were aimed to be identified by their highly conserved ITS region as well as studied in vanillinsupplemented medium through, vanillin-supplemented plates, TLC and Neubauer’s chamber to find out which, among the several isolates tested, were the most resistant ones, understand how they take up vanillin and how their growth is affected by the presence of the phenolic compound. Two strains were identified as Rhodotorula babjevae. One of them, L4, together with LS22, Rhodosporidium kratochvilovae, could withstand and biodegrade high concentrations of vanillin, producing biodegradation products with Rf values similar to the ones know for vanillic acid and vanillyl alcohol. Better growth in medium supplemented with small doses of vanillin was found, as well as disparity among the same species and their metabolic features, therefore, herbicides resistance was suggested as a reason for strains divergence. Further morphological-species comparison could also describe if there exist a relation between them.Estudios sobre la biodegradación de ligninas por levaduras basidiomicetes muestran la conversión de lignina en distintos productos de degradación, entre los cuales se encuentra la vainillina, un fuerte inhibidor de la fermentación y el crecimiento de levaduras. Las levaduras Sporidiobolales utilizadas en estos experimentos han intentado ser identificadas a través de la región ETI, muy conservada, además de estudiadas en medios suplementados con vainillina mediante placas suplementadas con vainillina, CCF y cámara de Neubauer para averiguar cuáles son las cepas más resistentes, entender cómo metabolizan la vainillina y cómo su crecimiento se ve afectado por la presencia de dicho compuesto. Dos cepas fueron identificadas como Rhodotorula babjevae. Una de ellas, L4, junto con con la cepa LS22, Rhodosporidium kratochvilovae, pudieron soportar y biodegradar elevadas concentraciones de vainillina, originando productos de biodegradación con valores de Rf similares a los del ácido vanílico y alcohol vanílico previamente conocidos. Se encontró un crecimiento mejor en medios suplementados con pequeñas dosis de vainillina además de una disparidad entre mismas especies y sus características metabólicas, así, herbicidas han sido sugeridos como una posible causa en dicha divergencia. Una futura comparación morfología-especie podrá describir si existe relación entre ambos.
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Fougère-Danezan, Marie. "Phylogénie moléculaire et morphologique des Detarieae résinifères (Leguminosae : Caesalpinioideae) : contribution à l'étude de l'histoire biogéographique des légumineuses." Thèse, Toulouse 3, 2005. http://hdl.handle.net/1866/6594.
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Förster, Frank. "Making the most of phylogeny: Unique adaptations in tardigrades and 216374 internal transcribed spacer 2 structures." Doctoral thesis, 2010. https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-51466.
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The phylum Tardigrada consists of about 1000 described species to date. The animals live in habitats within marine, freshwater and terrestrial ecosystems allover the world. Tardigrades are polyextremophiles. They are capable to resist extreme temperature, pressure or radiation. In the event of desiccation, tardigrades enter a so-called tun stage. The reason for their great tolerance capabilities against extreme environmental conditions is not discovered yet. Our Funcrypta project aims at finding answers to the question what mechanisms underlie these adaption capabilities particularly with regard to the species Milnesium tardigradum. The first part of this thesis describes the establishment of expressed sequence tags (ESTs) libraries for different stages of M. tardigradum. From proteomics data we bioinformatically identified 144 proteins with a known function and additionally 36 proteins which seemed to be specific for M. tardigradum. The generation of a comprehensive web-based database allows us to merge the proteome and transcriptome data. Therefore we created an annotation pipeline for the functional annotation of the protein and nucleotide sequences. Additionally, we clustered the obtained proteome dataset and identified some tardigrade-specific proteins (TSPs) which did not show homology to known proteins. Moreover, we examined the heat shock proteins of M. tardigradum and their different expression levels depending on the actual state of the animals. In further bioinformatical analyses of the whole data set, we discovered promising proteins and pathways which are described to be correlated with the stress tolerance, e.g. late embryogenesis abundant (LEA) proteins. Besides, we compared the tardigrades with nematodes, rotifers, yeast and man to identify shared and tardigrade specific stress pathways. An analysis of the 50 and 30 untranslated regions (UTRs) demonstrates a strong usage of stabilising motifs like the 15-lipoxygenase differentiation control element (15-LOX-DICE) but also reveals a lack of other common UTR motifs normally used, e.g. AU rich elements. The second part of this thesis focuses on the relatedness between several cryptic species within the tardigrade genus Paramacrobiotus. Therefore for the first time, we used the sequence-structure information of the internal transcribed spacer 2 (ITS2) as a phylogenetic marker in tardigrades. This allowed the description of three new species which were indistinguishable using morphological characters or common molecular markers like the 18S ribosomal ribonucleic acid (rRNA) or the Cytochrome c oxidase subunit I (COI). In a large in silico simulation study we also succeeded to show the benefit for the phylogenetic tree reconstruction by adding structure information to the ITS2 sequence. Next to the genus Paramacrobiotus we used the ITS2 to corroborate a monophyletic DO-group (Sphaeropleales) within the Chlorophyceae. Additionally we redesigned another comprehensive database—the ITS2 database resulting in a doubled number of sequence-structure pairs of the ITS2. In conclusion, this thesis shows the first insights (6 first author publications and 4 coauthor publications) into the reasons for the enormous adaption capabilities of tardigrades and offers a solution to the debate on the phylogenetic relatedness within the tardigrade genus ParamacrobiotusDer Tierstamm Tardigrada besteht aus derzeitig etwa 1000 beschriebenen Arten. Die Tiere leben in Habitaten in marinen, limnischen und terrestrischen Ökosystemen auf der ganzen Welt. Tardigraden sind polyextremophil. Sie können extremer Temperatur, Druck oder Strahlung widerstehen. Beim Austrocknen bilden sie ein so genanntes Tönnchenstadium. Der Grund für die hohe Toleranz gegenüber extremen Umweltbedingungen ist bis jetzt nicht aufgeklärt worden. Unser Funcrypta Projekt versucht Antworten darauf zu finden, was die hinter dieser Anpassungsfähigkeit liegenden Mechanismen sind. Dabei steht die Art Milnesium tardigradum im Mittelpunkt. Der erste Teil dieser Arbeit beschreibt die Etablierung einer expressed sequence tags (ESTs) Bibliothek für verschiedene Stadien von M. tardigradum. Aus unseren Proteomansatz konnten wir bislang 144 Proteine bioinformatisch identifizieren, denen eine Funktion zugeordnet werden konnte. Darüber hinaus wurden 36 Proteine gefunden, welche spezifisch für M. tardigradum zu sein scheinen. Die Erstellung einer umfassenden internetbasierenden Datenbank erlaubt uns die Verknüpfung der Proteom und Transkriptomdaten. Dafür wurde eine Annotations-Pipeline erstellt um den Sequenzen Funktionen zuordnen zu können. Außerdem wurden die erhaltenen Proteindaten von uns geclustert. Dabei konnten wir einige Tardigraden-spezifische Proteine (tardigrade-specific protein, TSP) identifizieren die keinerlei Homologie zu bekannten Proteinen zeigen. Außerdem untersuchten wir die Hitze-Schock-Proteine von M. tardigradum und deren differenzielle Expression in Abhängigkeit vom Stadium der Tiere. In weiteren bioinformatischen Analysen konnten wir viel versprechende Proteine und Stoffwechselwege entdecken für die beschrieben ist, dass sie mit Stressreaktionen in Verbindung stehen, beispielsweise late embryogenesis abundant (LEA) Proteine. Des Weiteren verglichen wir Tardigraden mit Nematoden, Rotatorien, Hefe und dem Menschen, um gemeinsame und Tardigraden-spezifische Stoffwechselwege identifizieren zu können. Analysen der 50 und 30 untranslatierten Bereiche zeigen eine verstärkte Nutzung von stabilisierenden Motiven, wie dem 15-lipoxygenase differentiation control element (LEA). Im Gegensatz dazu werden häufig benutzte Motive, wie beispielsweise AU-reiche Bereiche, gar nicht gefunden. Der zweite Teil der Doktorarbeit beschäftigt sich mit den Verwandtschaftsverhältnissen einiger kryptischer Arten in der Tardigradengattung Paramacrobiotus. Hierfür haben wir, zum ersten Mal in Tardigraden, die Sequenz-Struktur-Informationen der internal transcribed spacer 2 Region als phylogenetischen Marker verwendet. Dies erlaubte uns die Beschreibung von drei neuen Arten, welche mit klassischen morphologischen Merkmalen oder anderen molekularen Markern wie 18S ribosomaler RNA oder Cytochrome c oxidase subunit I (COI) nicht unterschieden werden konnten. In einer umfangreichen in silico Simulationsstudie zeigten wir den Vorteil der bei der Rekonstruktion phylogenetischer Bäume unter der Hinzunahme der Strukturinformationen zur Sequenz der ITS2 entsteht. ITS2 Sequenz-Struktur-Informationen wurden außerdem auch dazu benutzt, eine monophyletische DO-Gruppe (Sphaeropleales) in den Chlorophyceae zu bestätigen. Zusätzlich haben wir eine umfassende Datenbank, die ITS2-Datenbank, überarbeitet. Dadurch konnten die Sequenz-Struktur-Informationen verdoppelt werden, die in dieser Datenbank verfügbar sind. Die vorliegende Doktorarbeit zeigt erste Einblicke (6 Erstautor- und 4 Koautor-Publikationen) in die Ursachen für die hervorragende Anpassungsfähigkeit der Tardigraden und beschreibt die erfolgreiche Aufklärung der Verwandtschaftsverhältnisse in der Tardigradengattung Paramacrobiotus
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Huang, Liyin, and 黃莉媖. "Identification of Medically Relevant Molds by Sequence Analysis of the Internal Transcribed Spacer Regions 1 and 2." Thesis, 2003. http://ndltd.ncl.edu.tw/handle/73294799409112237403.
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碩士國立成功大學
醫學工程研究所碩博士班
91
The frequency of fungal infections has increased in recent years due to the increasing number of immunocompromised patients. Rapid identification of clinically relevant molds is useful for appropriate treatment with antifungal agents. The conventional methods for fungal identification mostly rely on morphological and biochemical tests; these methods are time-consuming and may be inaccurate. In this study, the feasibility of identification of medically important fungi by using the sequences of the internal transcribed spacer regions 1 and 2 (ITS1 and ITS2) was evaluated. The ITS data in GenBank were insufficient to identify clinically relevant fungi. The ITS1 and ITS2 regions from 198 reference strains (69 species) were amplified by PCR and sequenced; these sequences in combination with data in the GenBank were used to construct an ITS sequence database for the identification of molds. Totally 110 ITS1 and 108 ITS2 sequences were determined in this study. It was found that the ITS similarity scores among strains of the same species usually exceeded 0.95, whereas the scores were less than 0.95 among different species. The database was used to test 70 strains (30 species) of clinical isolates; a sensitivity of 89% and a specificity of 90% were obtained. It was concluded that the present method provides a rapid, simple, and reliable alternative to conventional methods for identification of medically important molds.
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Wei, Nu-Wei Vivian, and 魏汝薇. "Molecular evolution of the ribosomal internal transcribed spacer 2 in Acropora (Cnidaria; Scleractinia): effects of paralogy and ancestral polymorphism." Thesis, 2001. http://ndltd.ncl.edu.tw/handle/48853889364288556403.
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碩士國立臺灣大學
海洋研究所
89
Acropora is the most species-rich genus of scleractinian corals. Different Acropora species spawn synchronously and may increase possibility for interspecific hybridization. Some studies have suggest introgressive hybridization as an explanation for the reticulate evolution of Acropora, based on evidence from in vitro hybridization and molecular data. However factors other than hybridization, such as random sorting of ancestral polymorphism (lineage sorting), paralogy (gene duplication/ extinction), can also lead to a reticulate pattern of molecular phylogeny. In this study, population parameters were estimated for the ribosomal DNA internal transcribed spacer 2 (ITS2) for the three Acropora species A. muricata, A. hyacinthus and A. valida from two populations in Taiwan and compared with two geographically distant populations (Australia and Indonesia). A total of 119 Acropora ITS2 sequences from 35 individuals were used in the analyses. Analysis of molecular variance (AMOVA) indicated that most of the ITS2 variation occurred within populations, ranging from 85.87 % in A. muricata, to 91.50 % in A. hyacinthus, based on geographic populations. In contrast, based on the variants, most of the variation occurring among variants, ranging from 52.78 % in A. valida to 71.56 % in A. muricata. Phylogenetic analysis of ITS2 DNA sequences showed that ITS2 variants are dispersed among species in the phylogenetic tree, and do not correspond to geographic localities. It is suggested therefore that, instead of hybridization, the primary factors determining the evolutionary pattern of Acropora ITS2 is retention of paralogy and ancestral polymorphism. Thus, a reticulate evolutionary history of Acropora should be inferred with caution.
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Wang, Shiou-Wen, and 王秀文. "Internal Transcribed Spacer and Quick Identification of Botrytis elliptica." Thesis, 2000. http://ndltd.ncl.edu.tw/handle/42637919705770998864.
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碩士國立臺灣大學
植物病理學研究所
88
Botrytis spp. includes important plant pathogenic fungi which cause gray mold or leaf blight disease on several plants. Botrytis elliptica causes Botrytis blight on leaves and flowers of lily plants. B. gladiolorum mainly infects on gladiolus plants. The host range of B. cinerea includes plants of over 100 different species. In this study, the pathogenicity assay showed that all three Botrytis spp. could infect lily petal without wounding, but only B. elliptica could infect lily leaves without wounding. B. cinerea could only infect the wounding sites of lily leaves. B. gladiolorum could not infect lily leaves even with wounding. For the purpose of quick differentiation of B. elliptica, B. cinerea, and B. gladiolorum, PCR techniques was used to establish a rapid and sensitive procedure of detection. Primers are designed according to the sequence of ITS1 and ITS2 regions of Botrytis spp. Among four primer pairs, primer pair 102/106 (primer 102 designed from ITS1 of B. gladiolorum) could amplify a DNA fragment of predicted size from B. gladiolorum, but not from B. elliptica and B. cinerea. Primer pair 102-1/103 (primer 103 designed from ITS2 of B. elliptica and B. cinerea) could amplify a DNA fragment of predicted size from both of B. elliptica and B. cinerea, but not from B. gladiolorum. Both primer pairs could not amplify expected DNA fragments from Sclerotinia sclerotiorum. Primer pair 102-1/103-2 could amplify a DNA fragment of predicted size from Botrytis gladiolorum, but still obtained weak signal from B. elliptica, and B. cinerea. Primer pair 102-1/106 (designed from ITS conserved region of Botrytis spp.) could amplify DNA fragment of predicted size from B. elliptica, B. cinerea, and B. gladiolorum, and also from Sclerotinia sclerotiorum. All of four primer pairs could not generate detection signal from other pathogenic fungi of nine species and lily. Restriction map of ITS1-5.8S rDNA-ITS2 region of B. elliptica, B. cinerea, and B. gladiolorum were analyzed. PleI-digested DNA pattern of B. gladiolorum was different from that of B. eliptica and B. cinerea.
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Celio, Gail J. "Poinsettia powdery mildew immunolabeling, infection, and internal transcribed spacer regions /." 2002. http://purl.galileo.usg.edu/uga%5Fetd/celio%5Fgail%5Fj%5F200212%5Fphd.
Full text
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Lin, Shu-Chu, and 林淑珠. "Study on internal transcribed spacer sequences and antioxidative activities of Chlorella sp. and Arthrospira maxima." Thesis, 2002. http://ndltd.ncl.edu.tw/handle/08212331224589294017.
Full textAbstract:
碩士國立屏東科技大學
熱帶農業研究所
90
Using a light microscope to study Spirulina sp. or Chlorella sp., which were obtained from the Marine Products Research Institute at Tungkang, Pingting County, only its external morphology, as spiral or round shape, could be observed. However, when supplemented by a transmission electron microscope, its internal structures could be determined, showing septa, thylacoid and a not-well-defined nucleus in Spirulina sp. and cilia, pyrenoid and thylacoid in Chlorella sp. When comparing rDNA of Spirulina sp. and Chlorella sp. with those deposited in GenBank, the results showed that there was 100% similarity between Spirulina sp. and Arthrospira maxima (AJ292323) and about 71-76% similarity between Chlorella sp. and Nannochloris sp. The study on the ability of the free radical DPPH in eliminating antioxidation showed that, when comparing A. maxima and Chlorella sp. with the standards VitE and BHA under the same concentration of 0.4mg/ml, the antioxidative activities of these two algae were about half of that of VitE and BHA. The antioxidative activities were arranged on the basis of their intensity, in a descending order, VitE>BHA>Chlorella sp. >A. maxima. When the concentration was increased to 1.2 mg/ml, the antioxidative activities of A. maxima and Chlorella sp. increased to about 90%, which was close to that of VitE and BHA. That intensity was at its peak at this concentration and remained so in a stable condition. After purifying through the DEAE anion exchange column, the activities of the superoxide dismutase (SOD) of A. maxima. were 875U/mg, which was 18.3 times more than that of its initial extracts. The same activities of Chlorella sp. were 960U/mg, about 17.9 times of its initial extracts. This result showed that the SOD’s of these two algae were three times higher than that of Ganoderma lucidum and mushroom, demonstrating that there were high SOD activities in A. maxima and Chlorella sp. A. maxima and Chlorella sp., therefore, have a high potential for developing into pharmaceutical or health-related products, emphasizing as having specialized antioxidative characteristics.
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Chen, Chiao-Yi, and 陳巧宜. "Analysis of Ribosomal DNA Internal Transcribed Spacer Sequence and Secondary Metabolites of Metarhizium anisopliae Varieties." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/getw4y.
Full textAbstract:
碩士國立嘉義大學
生物資源學系研究所
97
Metarhizium anisopliae (Metsch.) Sorokin, a fungal entomopathogen, can infect more than 200 kinds of insect; the it is a value fungus for extensive studies both in industry and medicine. In the present study, 32 isolates of M. anisopliae collected from domestic and abroad were analyzed for ribosomal DNA internal transcribed spacer (rDNA-ITS) and secondary metabolites. The results of the difference between varieties were subjected the collected species. The difference of DNA sequence were analyzed by method to construct Neighbor-Joining (NJ) and Maximum-Parsimony (MP) phylogeny tree. The differences of secondary metabolites, mainly destruxins and cytochalasins of M. anisopliae varieties were also to be subjected to statistically analysis further. Ourrently. We use Ma0407, which collected from Nanto with analogs of cytochalasin, and BCRC 35507 purchased from Food Industry Research and Development Institute with destruxins. I employ Ma0407 and BCRC 35507 as reference group for chemodiversity study. After high-performance liquid chromatography (HPLC) analysis, the data reveals that ML13 and Ma0407 could produce the identical analogs of cytochalasins, in addition, BCRC 35507, Ma0319, BCRC 35520, and ARSEF 488, 7488, 5369 have the same destruxins; however, the classification and that quantity of destruxins in M. anisopliae strains are generally different, but destruxins A, B, B2/E2 and C exist commonly in every M. anisopliae isolate. In the present study, we explore the phylogeny of Metarhizium species and of Metarhizium varieties through rDNA-ITS sequencing method, and then investigate the chemodiversity of secondary metabolites between varieties. Results show that even if different M. anisopliae strains that possess identical rDNA-ITS sequence, meaning they have close phylogenetic relationship, cannot produce completely identical metabolites.
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hau, gou guo, and 勾國豪. "The analysis of internal transcribed spacer and its applications on the detection of orchid bacterial diseases." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/35210152903129477720.
Full textAbstract:
碩士中華醫事學院
生物科技研究所
93
There are three species of bacterial can infect the local Phalaenopsis, three of them are Pectobacterium chrysanthemi, Acidovorax avenae subsp. cattleyae and Burkholderia gladioli. In this study, we use the in-house bacterial internal transcribed spacer database to analysis the sequence between the bacterial that can infect the Phalaenopsi. We design primers AAC-ITS and BG-ITS based on the results of the analysis of Acidovorax avenae subsp. cattleyae and Burkholderia gladioli ITS sequence, and the primers pecS、pecM and idgA designed by indigoidine related gene, to detect the Pectobacterium chrysanthemi. The primers we use in this study reveals perfect results to their on response bacterial. In the sensitivity test, primer idgA can detect the Pectobacterium chrysanthemi with 1.2x104 cells, primer AAC-ITS can detect Acidovorax avenae subsp. cattleyae with1x102 cells and primer BG-ITS can detect Burkholderia gladioli with 1.2x104 cells. The primers designed in this study can obviously and accurately detect the bacterial that can infect the local Phalaenopsis.
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Abdennadher, Mourad. "Molecular fingerprinting, rDNA internal transcribed spacer sequence, and karyotype analysis of Ustilago hordei and related smut fungi." Thesis, 1995. http://hdl.handle.net/1957/34737.
Full textAbstract:
Inbreeding of the avirulent physiologic race 8 strains of Ustilago hordei was
purported to have increased its pathogenicity in a stepwise manner that led to a highly
pathogenic race, designated race 14. The analysis of electrophoretic karyotypes and
restriction fragment length polymorphism profiles detected with a telomere-specific probe
(TelomereRFLP) in races 8 and 14, and progeny obtained by selfing race 8 strains,
revealed substantially changed patterns in the purported progeny of the second generation
of selfing, and the race 14 strain. The telomereRFLP patterns in strains purported to be
the progeny of the second generation of inbreeding of race 8, were unlike both race 8 and
race 14 strains, and identical to those of U. hordei physiologic races 10 and 13. These
data suggest that the progeny believed to have been derived from the second selfing of
race 8 strains were clonal lineages from either race 10 or race 13 strains, rather than the products of meiosis of race 8 teliospores. The electrophoretic karyotypes resolved from
race 8 and 14, revealed chromosome-length polymorphisms (CLPs) that were similar in magnitude to those reported among strains of the fourteen physiologic races of U. hordei, rejecting the postulate that race 14 is a lineage derivative from race 8.
Electrophoretic karyotyping and ribosomal DNA (rDNA) sequence analysis of eight Ustilago species revealed that the four sporidium-forming species, U. avenae, U. hordei, U. kolleri, and U. nigra, form a coherent group. Ten probes detected a maximum of 15% CLPs among the putative homologous chromosomes of theses species, and the internal transcribed spacer (ITS) amplified from these species shared 97-99% sequence identity. The taxonomic distinctness of U. maydis from the rest of the smut fungi was evidenced by its divergence at 7 nucleotides in the 5.8S rDNA coding region.Graduation date: 1996
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Grubisha, Lisa C. "Systematics of the genus Rhizopogon inferred from nuclear ribosomal DNA large subunit and internal transcribed spacer sequences." Thesis, 1998. http://hdl.handle.net/1957/36754.
Full textAbstract:
Rhizopogon is a hypogeous fungal genus that forms ectomycorrhizae
with genera of the Pinaceae. The greatest number and species of
Rhizopogon are found in coniferous forests of the Pacific Northwestern
United States, where members of the Pinaceae are also concentrated.
Rhizopogon spp. are host-specific primarily with Pinus spp. and
Pseudotsuga spp. and thus are an important component of these forest
ecosystems. Rhizopogon includes over 100 species; however, the
systematics of Rhizopogon have not been well understood. Currently the
genus is placed in the Boletales, an order of ectomycorrhizal fungi that are
primarily epigeous and have a tubular hymenium. Suillus is a stipitate
genus closely related to Rhizopogon that is also in the Boletales and host
specific with Pinaceae. I examined the relationship of Rhizopogon to
Suillus and other genera in the Boletales. Infrageneric relationships in
Rhizopogon were also investigated to test current taxonomic hypotheses
and species concepts. Through phylogenetic analyses of large subunit and
internal transcribed spacer nuclear ribosomal DNA sequences, I found that
Rhizopogon and Suillus formed distinct monophyletic groups.
Rhizopogon was composed of four distinct groups; sections Amylopogon
and Villosuli were strongly supported monophyletic groups. Section
Rhizopogon was not monophyletic, and formed two distinct clades. Section
Fulviglebae formed a strongly supported group within section Villosuli.
Taxonomic revisions were proposed. Suillus, Truncocolumella, and the
Gomphidiaceae were transferred to the Rhizopogonaceae. In Rhizopogon,
sections Amylopogon, Rhizopogon, and Villosuli were elevated to
subgenera. Subgenus Roseoli was erected to accommodate the second
section Rhizopogon Glade. In section Fulviglebae, Stirps Vinicolor,
Rhizopogon ochraceisporus, R. subclavitisporus, and R. clavitisporus were
transferred to subgenus Villosuli while the remaining species in section
Fulviglebae were transferred to subgenus Rhizopogon.Graduation date: 1999
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Procházková, Kateřina. "Diverzita a druhový koncept u komplexu Vischeria/Eustigmatos (Eustigmatophyceae)." Master's thesis, 2012. http://www.nusl.cz/ntk/nusl-305377.
Full textAbstract:
Vischeria and Eustigmatos are closely related genera occurring in terrestrial habitats. These genera were distinguished by the differences in the features of the cell wall (projections and ridges of different form, smooth surface respectively). Up to date three species of the genus Eustigmatos and twelve species of the genus Vischeria have been described, but nine of the Vischeria species have been rarely, if ever, observed since the original description. This work is focused on evaluating molecular variability, diversity, and taxonomy of the Vischeria/Eustigmatos complex. Ninety seven strains, obtained from public algal collections or newly isolated from localities from all over the world, were studied, including the type strains of two Eustigmatos species and three Vischeria species. Phylogenetic analyses of the ITS2 rDNA and rbcL sequences showed that these genera are not genetically separated. The five types strains each represented a separate evolutionary lineage. Some of the additional lineages included strains morphologically corresponding to the species Eustigmatos magnus. Some of the newly isolated strains are according to the markers examined genetically indistinguishable from known strains from public algal collections. However, some of them are new lineages. Only one of the phylogenetic...
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Leaw, Shiang-Ning, and 劉向寧. "Identification of Medically Important Yeasts by Sequence Analysis of the Internal Transcribed Spacer Regions and by an Oligonucleotide Array." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/42523302969995742423.
Full textAbstract:
博士國立成功大學
醫學工程研究所碩博士班
95
Infections caused by yeasts have increased in recent decades due to the increasing population of immunocompromised patients. In addition to Candida spp., infections caused by less common species such as Pichia, Rhodotorula, Trichosporon, and Saccharomyces have been widely reported. Accurate and rapid identification of yeast pathogens is important for appropriate treatment of patients with antifungal agents. The objectives of this study were to evaluate the feasibility of using the sequence of ribosomal DNA internal transcribed spacer (ITS) regions and an oligonucleotide array to identify clinically important yeasts. The first part of this thesis evaluated the feasibility of ITS sequencing for yeast identification. Both ITS1 and ITS2 regions of 373 strains (86 species) including 299 reference strains and 74 clinical isolates were amplified by PCR and sequenced. The sequences were compared to reference data available at the GenBank database using BLAST (Basic Local Alignment Search Tool) for species determination. The rates of correct identification by ITS1 and ITS2 sequence analysis were 96.8% (361/373) and 99.7% (372/373), respectively. The second part of this thesis was to evaluate primer labeling (one or both primers) and the use of sense or antisense probe on the hybridization signal. It was found that labeling of both primers produced, under most conditions, stronger hybridization signal. However, the effect of using sense or antisense probe on hybridization signal was not predictable. The third part of this thesis was to develop an oligonucleotide array to identify 77 species (16 genera) of clinically relevant yeasts based on the ITS sequence. The ITS regions were amplified by PCR with a pair of fungus-specific primers, followed by hybridization of the digoxigenin-labeled PCR product to a panel of oligonucleotide probes immobilized on nylon membrane for species identification. A collection of 452 yeast strains (419 target and 33 nontarget strains) was tested, and a sensitivity of 100% and a specificity of 97% were obtained by the array. The detection limit of the array was 10 pg of yeast genomic DNA per assay. In addition, 42 yeast-positive blood cultures were analyzed by the oligonucleotide array, and all specimens were correctly identified. In conclusion, yeast identification by both methods (sequence analysis and oligonucleotide array) is highly reliable and can be used as an alternative to the conventional identification methods. The whole procedure can be finished within 24 h starting from isolated colonies.
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Lin, Hsueh-Fang, and 林雪芳. "Identification of insect cell lines and cell line cross-contamination (CLCC) by internal transcribed spacer region of ribosomal DNA." Thesis, 2008. http://ndltd.ncl.edu.tw/handle/94004670050695777094.
Full textAbstract:
碩士國立臺灣大學
昆蟲學研究所
96
Insect cell lines are useful tools in the study of virus basic biology and production of recombinant proteins, especially by baculovirus expression vector system (BEVS). In most cell culture laboratories, several different cell lines are routinely used, maintained, or handled, these cell lines should be regularly monitored to avoid cell line cross-contamination (CLCC). We developed a technique for identification of species origin and CLCC detection for insect cell lines (NTU-LY, NTU-PN, IPLB-LD652Y, and Sf9 cell lines) based on the sequence of internal transcribed spacer (ITS) of ribosomal DNA (rDNA). The identity of the amplicon’s sequences between insect and its homologous cell line was up to 95 %~99 %, thus these amplicon’s sequences could be used to confirm the species origin of a cell line. Two endonucleases, PstI and HindII based on these sequences of amplicons, were selected for rapid identification of insect cell lines by PCR-RFLP (PCR-restriction fragment length polymorphism). The homologous cell lines could be examined by RAPD-PCR (random amplified polymorphic DNA – PCR) method with a selected oligonucleotide primer, OPU-10. Three species-specific primer sets, Ly-ITS1/Ly-ITS2, ITS1-1/Ld-ITS1, and Sf9-F2/ITS4, were then designated for identification of LY, LD, and Sf9 cells, respectively, by SS-PCR (species-specific PCR). Therefore three methods, RFLP-PCR, RAPD and SS-PCR, can be used to define cell origin and avoid CLCC. Otherwise, the mass production of NTU-LY cells was also undertaken by suspension culture after confirmations of cell origin and free from CLCC. The optimal condition of LY cell suspension culture was 1 x 106 cells/ml, in 200 ml culture volume and 65 RPM at 28℃. The LyxyNPV OBs (occlusion bodies) production reached to 70.5 ~ 97 OBs/cell that was much higher than monolayer infection (27.7 OBs/cell).
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Chang, Tsong-Chih, and 張聰智. "Analysis of genetic diversity of Ganoderma species based on ribosomal DNA internal transcribed spacer (ITS) region sequences and laccases gene." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/88901216809281292868.
Full textAbstract:
碩士國立中興大學
生命科學院碩士在職專班
99
With the rapid development of molecular biology techniques, the taxonomy of fungi has been transformed from by morphological phenotype into a new era of molecular classification. In this study, we use of polymerase chain reaction (PCR) technology to amplify both the consensus sequence of the laccase gene and the ribosomal DNA internal transcribed spacer sequences (ITS) of 15 Ganoderma strains (including four BCRC standard strains and 11 of wild type isolates) and three exceptionally genus strains. The specific 595~915 bp of PCR product of TS1-ITS2 fragement and 1401~1651 bp of laccase gene fragments have been amplified, then cloning by TA- strategy for DNA sequencing. Each DNA sequence has been applied into NCBI database for sequence alignment by similarity analysis to compare the naming of molecular identification and traditional morphological classification. And, respectively, alignment by CLUSTAL program to establish the phylogenetic tree.
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Wu, Kun-Chan, and 吳坤禪. "Genetic identification of inter- and intra-species of chicken Eimeria in Taiwan by the second internal transcribed spacer ribosomal DNA." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/46203084432217993404.
Full textAbstract:
碩士國立屏東科技大學
生物科技研究所
95
Coccidiosis in chickens caused by protozoan parasites of Eimeria mainly infects epidermal cells of gastro-intestinal tracts, causing acute and chronic enteritis as well as bloody stool. These result in growth tardiness, increased rates of sensitivity to other diseases, and indirectly increased death rates of animals. It is, thus, a protozoan disease that causes great severity and high economic losses. Using traditional methods in diagnosis, the results may be incorrect due to multiple similarities in morphologies and co-infections of different Eimeria species. Recent development of polymerase chain reaction (PCR) has been considered as a more stable and correct technology compared to the traditional methods. The aims of this study were to identify species of Eimeria by both traditional and molecular biological methods, and further establish a data base based on the species characterization. The methods include cloning of one single oocyte that was microinjected into chickens. So far, the identification of 4 Taiwanese species, E. acervulina, E. maxima, E. mitis, and E. tenella has been completed based on the morphological characterization. This thesis describes the use of internal transcribed spacer-2- (ITS-2-) specific primers followed by PCR for the identification of these 4 Eimeria species. These PCR products were further digested with 3 restriction endonucleases, HhaI, RsaI and TaqI and analyzed using restriction fragment length polymorphisms (RFLP) by agarose gel electrophoresis. The electrophoretic data show that the sizes of PCR-RFLP were 440 base pairs (bp) for E. acervulina, 370 and 580 bp for E. maxima, 450 and 580 bp E. mitis, and 580 bp for E. tenella. Sequence alignments between nucleotides of E. acervulina, E. maxima, and E. tenella and those published in GenBank showed 94~99%, 78~96%, and 99~100% homology, respectively. Among these sequences, the 580-bp fragments shown in E. maxima and E. mitis had 99% homology to that of the 580-bp fragment in E. tenella. The nucleotide position on 151 for E. tenella and E. maxima was guanine (G) but thymidine (T) for E. mitis; and on the 493 position cytosine (C) for E. maxima and E. mitis, whilst T for E. tenella. Data from sequence alignments among these 4 Eimeria ITS-2 showed 52~67% homology in their nucleotides. Based on our knowledge, these has been no published data for the ITS-2 sequences of E. mitis, this report may represent the novel data in this species of Eimeria.
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Hsu, Hsi-Shih, and 徐希世. "Studies of the morphological taxonomy and phylogenetic analysis using internal transcribed spacer of ribosomal DNA on the genus Marasmius in Taiwan." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/30286905061370194892.
Full textAbstract:
博士國立中興大學
生命科學系
94
The morphological taxonomy and phylogenetic analysis of Marasmius were studied. In experimental period, forty-nine specimens were collected from Nantou Hui-Sun forest, Nantou Ming-Tan hydraulic power plant, Nantou Guan-Yin waterfalls, Nantou Lien-Hua-Chih forest, Kaohsiung Cheng-Ching Lake and Hsinchu Niao-Zui Mountain. Seven species were identified including Marasmius androsaceus, M. candidus, M. caricis, M. crinis-equi, M. maximus, M. rhyssophylus and M. siccus. There are two new records for Taiwan namely M. caricis and M. rhyssophylus. To study the relationship in the genus Marasmius, internal transcribed spacer（ITS）1 and 2 regions of rDNA were amplified by PCR. Forty DNA sequences of ITS were compared by using ClustalW program and grouped by principle component analysis (PCA). Phylogenetic tree was drawn by bootstrap method. A key of Marasmius established by using internal transcribed spacer DNA sequence.
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Huang, Tzu-pi, and 黃姿碧. "Application of rDNA internal transcribed spacer (ITS) sequence characteristics as an aid for the identification and phylogenetic study of Colletotrichum spp." Thesis, 1999. http://ndltd.ncl.edu.tw/handle/82688830118490959818.
Full textAbstract:
碩士國立中興大學
植物病理學系
87
Colletotrichum gloeosporioides (Cg), the causal agent of anthracnose on various important crops, is the most notorious post-harvest pathogens greatly deteriorating the quality and shelf-life of agricultural products. The traditional way for identification of Colletotrichum spp. (C-spp) is quite often frustrating and time consuming mainly due to the wide host range of the pathogen, the great morphological variation; and what even worse was that cross infection might occur among different Colletotrichum spp. To cope with the increasing need of rapid detection and identification of this fungal pathogen from various agricultural products after the country become a member of the World Trade Organization (WTO), the potential application of rDNA ITS sequence characteristics as a biochemical tool was explored. The full length ITS rDNAs of a total of 29 C-spp isolates, and 7 non-C-sp isloates, were produced by polymerase chain reaction (PCR) amplification using the universal primer pair ITS1/ITS4 (White et al.1990). The Colletotrichum species tested included C. gloeosporioides (Cg), C. acutatum (Ca), C. musae (Cm), C. graminicola (Cgr), C. coccodes (Cco), C. capsici (Cc), C. dematium (Cd), C. lindemuthianum (Cl), and Colletotrichum sp. (C-sp) that species status awaited to be identified. The non- Colletotrichum species tested included Rhizopus oryzae, Penicillium digitatum, Pestalotia sp., Fusarium solani, Alternaria brassica, Ustilago esculenta, and Trichoderma reesici. The 600 bps amplicons were generated from all C-spp isolates tested as was expected; the amplicons from 7 non-C-spp isolates ranged from 500-800bps. Fifteen of the C-spp amplicons which include 7 from Cg; 2 each from Cc, Cgr; and one each from Cm, Cd, Cl and C-sp, were cloned and sequenced. The full length of these amplicons ranged from 535 to 555 bps; in which ITS1 region was from 160 to 181bps, ITS2 region was from 152 to 153bps; whereas the 5.8S rDNAs all appeared to be 160bps. Comparison of the inter-specific sequence identity indicated the existence of much greater divergence among ITS1 (identity reached only 53.9%) as compared to that among ITS2 region (identity was approximately 81.8%). Comparative analysis of the ITS full length and the ITS1 sequence data by the distance matrix method and the parsimony method both concluded these 15 tested isolates into 5 distinctive species groups namely, CG (C. gloeosporioides), CA (C. acutatum), CM (C. musae), CL (C. lindemuthianum), and CC (C. capsici). The sequence data also revealed that the ITS1 sequence of tested CG, CA and CC isolates were 99-100% identical to each compared isolate available in the GenBank/EMBL data bank; whereas for ITS2 region, only those from CA and CG isolates were greater than 98% in identity. For the rapid detection and identification of Colletotrichum spp. by PCR, the efficacy of the species specific primer pairs developed by Mills et al. (1992) were examined; the results appeared to be non-satisfactory. To resolve the problem, the heteroduplex mobility assay (HMA) was attempted using the ITS-rDNA amplicons of all 29 tested C-spp isolates as the targets. The analysis by the above described distance matrix method using the distance value estimated from the mobility changes concluded the 29 isolates into 5 distinctive species groups same as that by sequence data analysis. Moreover, from the repeatable band distribution characteristics, a total of 6 heteroduplex patterns (HP) were established. By the established HP fingerprint, the species group CG can be further divided into CG1 and CG2; while the other 4 species groups remain unchanged. In the established species group system, Cgr1, Cgr2 and Cd1 isolates, originally identified as C. graminicola and C. dematium respectively, were reclassified as member of CC since their ITS sequences were 98-100% identical to that of C. capsici. Likewise, the species status of Cg8 isolate was changed from C. musae to a member of CG; Cg2, Ca1, Cg9 and C-sp1 isolates were changed from C. gloeosporioides to be members of CA; and Cc1 isolate was changed from C. capsici to a member of CG2. Among these isolates that original identification were disputable, the species status of Cc1 was reexamined by the original author lately and proved the previous assignment as C. capsici might be a misinterpretation. The results discussed in this investigation, although only limited numbers of fungal isolates had been explored, appeared to provide evidence that sequence analysis of ITS rDNAs a dependable tool in the species level identification of Colletotrichum spp. Also the combination of the genus- or species-specific PCR together with HMA and HP analysis is easy to process, not time consuming, and provides repeatable and distinct data bases useful for species identification. The use of the established techniques for practical application in rapid detection and pathogen identification of anthracnose, and even other fungal diseases, are thus recommended.
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Ballard, Harvey E. "Phylogenetic relationships and infrageneric groups in Viola (Violaceae) based on morphology, chromosome numbers, natural hybridization and internal transcribed spacer (ITS) sequences." 1996. http://catalog.hathitrust.org/api/volumes/oclc/36205086.html.
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"Molecular phylogenetics of the Magnoliaceae: An analysis of the chloroplastpsbA gene, mitochondrial cox II gene and nuclear internal transcribed spacer region." Tulane University, 1997.
Find full textAbstract:
The family Magnoliaceae is recognized as a group composed of some of the most primitive extant angiospews, with a long evolutionary history. An extensive fossil record supports the antiquity of this plant family. The Magnoliaceae, as a component of the order Magnoliales, is an important group in classification systems. While several conflicting morphological studies have been undertaken to classify and clarify relationships within the family, there have been no molecular evolutionary studies to help provide a more coherent understanding of taxonomy within the Magnoliaceae To help elucidate the phylogenetic position of the Magnoliaceae relative to other angiosperms, I constructed a phylogenetic tree of 22 taxa using the chloroplast psbA gene. Only two most parsimonious trees were revealed using a maximum parsimony method (PAUP). The phylogenetic analysis suggested a bifurcation of taxa with the Magnoliales, Piperales, Laurales, and Illiciales a distinct and separate group from the more advanced agricultural plants Nuclear ribosomal internal transcribed spacer sequences (ITS1 and ITS2) were then used to provide a highly resolved classification for several Magnolia sections. Magnolia section Rytidospermum (nine species) is a paraphyletic group which was initially characterized based on two morphological features: (1) large whorl-like leaf arrangement and (2) a wrinkled inner seed coat, a trait not shared by all members. ITS sequence data shows that M. macrophylla, M. ashei, and M. dealbata form a distinct monophyletic group, separate from M. tripetala, M. hypoleuca, M. pyramidata, and M. fraseri (section Rytidospermum) Congruent molecular data from subunit II of cytochrome c oxidase (cox II) supports the separation of M. macrophylla, M. ashei, and M. dealbata from the other members of section Rytidospermum. The complete loss of the cox II intron from section Rytidospermum and the presence of the intron in the rest of the family (including the M. macrophylla group) provides a useful diagnostic marker for determining interspecific relationships in some members of the familyacase@tulane.edu
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Huang, Chun-Sheng, and 黃春申. "Differential Diagnosis and Analysis of Avian Eimeria Species by PCR using the Primers Derived from Internal Transcribed Spacer 1 (ITS 1)." Thesis, 2004. http://ndltd.ncl.edu.tw/handle/05340738952156226799.
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李偉基. "Detection and identification of pathogenic escherichia coli by using multiplex polymerase chain reaction and analysis of ITS(internal transcribed spacer)nucleotide sequences." Thesis, 2003. http://ndltd.ncl.edu.tw/handle/89845605087225624464.
Full textAbstract:
碩士國立屏東科技大學
獸醫學系
91
The final goal of this study is for the fast diagnosis of different types of pathogenic Escherichria coli including EHEC ( Enterohemorragic E.coli )、ETEC ( Enterotoxigenic E.coli )、EIEC ( Enteroinvasive E.coli )、 EPEC ( Enteropathogenic E.coli ) and EAEC ( Enteroaggregative E.coli ). Moreover , the target genes employed for the differentiation of these kind of pathogenic E. coli are VTⅠ, VTⅡ, ST, LT, ial , pCVD432, eaeAgen , H7 and uidA. However, the resulting data could provide with that the pathogenic E. coli can be diagnosed by using three tubes-one step multiplex PCR ( polymerase chain reaction ). The diagnostic process just took as short as two and half hours in terms of fast- diagnosis. The ITS ( Internal transcribed spacer ) region of DNA fragment was cloned and sequenced from EHEC, ETEC, EPEC, EIEC and EAEC respectively. After a phylogenic analysis by using DNA star softwire, three groups of distinct characteristics of E. coli were demonstrated as groupⅠ: ETEC and EHEC were characterized by secreting their specific enterotoxins or cytotoxins in clinics; group Ⅱ: EAEC alone was not found in humans other than animals; group Ⅲ: EIEC and EPEC: the pathogenesis of this group of E. coli was not proposed by the mechanism of toxin-producing. The resulting data showed that 10% divergence in DNA sequences between groupⅠand groupⅡ, 41.5% divergence between groupⅠand groupⅢand the same result as shown between groupⅡand groupⅢ. However, The sequences of ITS could be used for differentiation of different types of pathogenic E. coli based on phylogenic analysis and also coincided with the characteristics of the different pathogenic E. coli which are grouped described above.
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Harrower, Emma. "Using Barcode Similarity Groups to Organize Cortinarius Sequences." Thesis, 2010. http://hdl.handle.net/1807/25613.
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To improve fungal identification using a single DNA sequence, I introduce the Barcode Similarity Group (BSG) defined as a cluster of sequences that share greater than or equal to a threshold amount of genetic similarity with each other. As a test case, I created 393 BSGs from 2463 Cortinarius ITS sequences using a 94% similarity cut-off value in DOTUR. Some BSGs may contain multiple species. The BSG database was used to label environmental sequences, find misidentified or mislabeled sequences, and find potential cryptic species and novel species. Expert taxonomists will be needed to perform detailed morphological and phylogenetic studies to identify the individual species within each BSG. The main advantage of using BSGs is that it clusters together sequences using total genetic relatedness and does not rely on any taxonomy for identification. A website was created where the RDP Classifier is used to classify a query sequence into a BSG.
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Tsai, Fang-Mei, and 蔡芳媚. "Differential diagnosis of five avian Eimeria species by polymerase chain reaction using the primers derived from internal transcribed spacer 1 (ITS 1) sequence." Thesis, 2002. http://ndltd.ncl.edu.tw/handle/21764820910894222131.
Full textAbstract:
碩士國立臺灣大學
獸醫學研究所
90
Chicken coccidia is a protozoan parasite of the genus Eimeria. It is economically important that causes loses in the poultry industry globally. The various species can be distinguished on the basis of morphology of the oocyst and the location of parasite within intestinal cells, but these criteria could be unreliable. Arbitrarily-primed PCR has been utilized to fingerprint avian Eimeria species, but this method does not always give reproducible results due to low specificity. For this reason, PCR assays have been employed for the detection of Eimeria species in this study. Five sets of primer derived from the internal transcribed spacer 1 (ITS-1) regions were used to distinguish the five Eimeria species including Eimeria tenella, E. necatrix, E. brunetti, E. acervulina and E. maxima. We use vaccine as template for the five species. The similarities of five species sequences between the vaccine and Genbank are 94%~100%. Analysis for E. tenella ITS-1 partial sequence of Taiwan and Genbank, the similarity is 99.6%. PCR sensitivity test of E. tenella in Taiwan is 50 oocysts. Five sets of primer won''''t amplify any non-specific band of chicken genome or intestinal contents.
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Lin, Wen-Yu, and 林文煜. "Identification of three Bupleurum species through a rapid detection method using the sequence-specific oligonucleotide within ribosomal DNA internal transcribed spacer as probe." Thesis, 2004. http://ndltd.ncl.edu.tw/handle/9939xm.
Full textAbstract:
碩士國立清華大學
生物技術研究所
92
Authentication of Chinese medicinal herbs gains more difficulties with the limitation of few unique morphological characters, and the process after harvesting. This study aims to develop an efficient method for rapid identification of Chinese medicinal herbs based on molecular biotechnology. Bupleuri Radix is a valuable and well known crude drug of Chinese medicine that is widely used in health care and remedy of liver disease. This herb is also one of the commonly misused crude drugs in Taiwan. The first part of this thesis is to analyze the genetic divergence among Bupleurum samples using the ribosomal DNA internal transcribed spacer (rDNA ITS) as marker. Our results showed that the ITS sequences of the three examined Bupleurum species share high identity with only few variations. In the second part, a rapid detection method was developed for identification of B. kaoi Liu Chao et Chuang, B. falcatum and B. chinense DC. based on oligonucleotide array. Oligomers of 15-20 bases within ITS containing one to three variation sites among different species were bound to nylon membrane. The ITS fragments were amplified with PCR using Dig-11-dUTP labeling and hybridized to a membrane holding the designed probes. The intensity of fluorescence signal was used to distinguish the binding strength between the amplified target sequence and the probe on membrane. Subsequently the suspected Bupleurum can be precisely identified through evaluation of the hybridization result in a short time and low-price. The appropriate hybridization temperature was measured below the Tm value of probe about 10-13ºC in our experiments. The signals of eight SSOP sets can be acquired after single reaction when we set hybridization temperature at 40ºC. The hybridization results of signals of forty-eight SSOPs with six dried Bupleurum samples displayed high accuracy (more than 90%). This method is reliable for authentication on condition that the factors of detection are optimized and sufficient SSOPs from the adulterate Chinese medicinal herbs are designed.