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Gray, Stephen James. "The genotyping of Neisseria meningitidis by restriction fragment length polymorphism (RFLP) analysis." Thesis, Manchester Metropolitan University, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.360085.
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Spengler, C. J. "PCR-RFLP typification of microbes used in the production of a fermented fish product." Thesis, Stellenbosch : Stellenbosch University, 2001. http://hdl.handle.net/10019.1/52397.
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Thesis (MScFoodSc)--Stellenbosch University, 2001.ENGLISH ABSTRACT: The preservation of various fresh fish products is achieved by either smoking, salting, canning, freezing or fermenting a highly perishable raw product. Since many of these facilities are not readily available, the use of fermentation as a means of preserving the product has been extensively practiced. However, the fermentation of fish is a time consuming practise and only by accelerating the process would it be possible to ensure the production of a more cost effective and readily available safe end-product. The quality of the fermented fish product is partially determined by the fermentation conditions and the metabolic activity of the microbes present. The rapid identification of the microbes present during the fermentation would enable the selection of possible starters to ensure an accelerated production of high quality fermented fish products. This study was thus undertaken to develop identification fingerprints for bacteria isolated from fermented fish products. A 1300 bp fragment of the 16S rRNA genes of each of the bacteria previously isolated was successfully amplified using the PCR technique. The isolates included strains of the genera Bacillus, Staphylococcus, Sphingomonas, Kocuria, Brevibacillus, Cryseomonas, Vibrio, Stenotrophomonas and Agrobacterium. The data obtained can, therefore, be used in the identification of these microbes isolated from other similar fermented fish products. The fingerprints could also be used to assist in determining the dominant microbial populations responsible for the characteristic qualitative changes occurring in the fish product during fermentation. The microbial composition of a fermenting fish product partially determines the quality of the end-product, therefore, the use of selected bacterial starters could result in the accelerated production of a microbial safe fermented fish product. A further objective of this study was to accelerate the production of a fermented fish product by inoculating macerated trout with either selected lactic acid bacteria (LAB) or with selected bacteria with high proteolytic activity over a 30 day fermentation period. The LAB included a combination of Lactobacillus plantarum, Lactococcus diacetylactis and Pediococcus cerevisiae strains, whereas the bacteria with high proteolytic activity included strains of Kocuria varians, Bacillus subtilis, two strains of B. amyloliquefaciens and a combination of these bacterial species. The quality of the fermented product was determined using changes in product pH, titratable acidity (%TA) and free amino nitrogen (FAN) formation as efficiency parameters. The data obtained during the fermentation of the macerated trout showed that the selected starters did not have a significant effect on the pH decrease in the product over a 30 day fermentation period. The LAB strains did not have a significant effect on the %TA of the fermenting fish product, yet the presence of these bacteria appeared to limit the FAN production in the product. The bacteria with high proteolytic activity resulted in slightly enhanced %TA values and a higher FAN content in the fermented product. It was also determined that the LAB and Kocuria varians, in contrast to the Bacillus spp. inoculums, did not survive the fermentation conditions well, possibly due to the low pH environment. The presence of the starter bacteria in the fermenting fish mixture at the end of the fermentation was also successfully determined with the use of the PCR-RFLP technique. The fermented fish product, obtained at the end of the fermentation period, had a good aroma and compared favourably to similar commercially available fermented fish products. The use of different microbial starters could in future enable the production of a diverse range of high quality products, which could be produced and marketed locally.
AFRIKAANSE OPSOMMING: Die preservering van ‘n verskeidenheid vars vis produkte word bereik deur die hoogs bederfbare produk te rook, te sout, te blik, te vries of te fermenteer. Aangesien baie van hierdie fasiliteite nie geredelik beskikbaar is nie, is die gebruik van fermentasie as ‘n preserverings metode al ekstensief beoefen. Die fermentasie van vis is egter 'n tydsame proses en slegs deur die versnelling van die proses sal dit moontlik wees om die produksie van ‘n meer koste effektiewe en geredelike beskikbare veilige eindproduk te verseker. Die kwaliteit van die gefermenteerde vis produk word gedeeltelik bepaal deur die fermentasie kondisies en die metaboliese aktiwiteit van die mikrobes teenwoordig. Die vinnige identifikasie van die mikrobes teenwoordig gedurende die fermentasie sal die seleksie van moontlike suursels om die versnelde produksie van hoë kwaliteit gefermenteerde vis produkte moontlik maak. Hierdie studie is dus onderneem om identifikasie vingerafdrukke vir bakteriee wat gei'soleer is van gefermenteerde vis produkte moontlik te maak. ‘n 1300 bp fragment van die 16S rRNA gene van elkeen van die bakteriee wat voorheen gei'soleer is, is suksesvol geamplifiseer deur die PCR tegniek. Die isolate sluit in stamme van die genera Bacillus, Staphylococcus, Sphingomonas, Kocuria, Brevibacillus, Cryseomonas, Vibrio, Stenotrophomonas en Agrobacterium. Die data kan dus gebruik word in die identifikasie van hierdie mikrobes as dit gei'soleer word van ander gefermenteerde vis produkte. Die vingerafdrukke kan ook gebruik word om die dominante mikrobiese populasies wat verantwoordelik is vir die kenmerklike kwalitatiewe veranderinge wat plaasvind in die vis produk gedurende die fermentasie, te identifiseer. Die mikrobiese samestelling van ‘n fermenterende vis produk bepaal gedeeltelik die kwaliteit van die eindproduk, daarom kan die gebruik van geselekteerde bakteriese suursels die versnelde produksie van ‘n mikrobies veilige gefermenteerde vis produk teweeg bring. ‘n Verdere doel van hierdie studie was om die produksie van ‘n gefermenteerde vis produk te versnel deur fyngemaakte forel met of geselekteerde melksuurbakteriee of met geselekteerde bakteriee met hoë proteolitiese aktiwiteit te inokuleer oor ‘n 30 dag fermentasie periode. Die melksuurbakteriee het ingesluit ‘n kombinasie van Lactobacillus plantarum, Lactococcus diacetylactis en Pediococcus cerevisiae, terwyl die bakterieë met hoë proteolitiese aktiwiteit stamme van Kocuria varians, Bacillus subtilis, twee stamme van Bacillus amyloliquefaciens en ‘n kombinasie van hierdie bakteriese stamme ingesluit het. Die kwaliteit van die gefermenteerde produk is bepaal deur die veranderinge in die pH, titreerbare suur (%TS) en vrye amino stikstof (VAS) vorming van die produk as effektiwiteits parameters te gebruik. Die data wat verkry is gedurende die fermentasie van die fyngemaakte forel het gedui daarop dat die geselekteerde suursels nie ‘n merkwaardige effek op die afname in pH in die produk oor ‘n 30 dag fermentasie periode het nie. Die melksuurbakteriee het nie ‘n merkwaardige effek op die %TS van die gefermenteerde vis produk gehad nie, terwyl dit geblyk het dat die teenwoordigheid van hierdie bakterieë die produksie van VAS in die produk belemmer het. Die bakteriee met hoe proteolitiese aktiwiteit het ‘n effense verhoogde %TS en ‘n hoër VAS inhoud in die gefermenteerde produk veroorsaak. Dit is ook bepaal dat die melksuurbakteriee en Kocuria varians, in teenstelling met die Bacillus spp. inokulums, nie die fermentasie kondisies goed oorleef het nie, moontlik as gevolg van die lae pH omgewing. Die teenwoordigheid van die suursel bakteriee in die fermenterende vis mengsel aan die einde van die fermentasie is ook suksesvol bepaal met die PKR-RFLP tegniek. Die gefermenteerde vis produk, verkry aan die einde van die fermentasie periode, het ‘n goeie aroma gehad en het goed vergelyk met soortgelyke kommersieel beskikbare gefermenteerde vis produkte. Die gebruik van verskillende mikrobiese suursels kan in die toekoms die produksie van ‘n diverse reeks hoë kwaliteit produkte wat plaaslik geproduseer en bemark kan word moontlik maak.
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Jaffe, Benjamin. "Genome analysis of Hordeum bulbosum L. and hybrids with H. vulgare L." Thesis, University of Reading, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.302327.
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Carter, Bradley Robert. "IS1002-associated RFLP fingerprinting of Bordetella pertussis clinical isolates, as a means of strain typing and epidemiological analysis." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2001. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/MQ59367.pdf.
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Finnegan, Anna Kathryn. "The genetic structuring of Atlantic salmon (Salmo salar L.) populations in northwest Europe as revealed through nuclear microsatellite and mtDNA PCR-RFLP analysis." Thesis, University of Exeter, 2009. http://hdl.handle.net/10036/72213.
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The structuring of Atlantic salmon (Salmo salar L.) into discrete, genetically differentiated populations both within and between river catchments is well documented. The utilisation of this knowledge has proved valuable in a variety of evolutionary, ecological, managerial and conservation contexts. In this thesis, the genetic structuring of Atlantic salmon populations in northwest Europe was assessed in two catchments of very different sizes, using a range of molecular and associated population genetic methods; findings from the catchment level research are set in context by a broader phylogeographic study of post-glacial colonisation of the region. A regional study into the glacial origins and post-glacial colonisation routes of Atlantic salmon in northwest Europe was explored by analysing a pre-existing microsatellite dataset and supplementing it with haplotype data from mtDNA PCR-RFLP analysis of the same samples (N=702). Evidence from allele permutation tests undertaken on the microsatellite data alongside mtDNA haplotype frequencies suggested that there was a cryptic northern refuge in northwest France, with colonisation of the British Isles and Ireland occurring from this and the long-known Iberian Peninsula refuge. Catchment level studies were undertaken on the river Dart and river Tweed, involving 1151 fish being genotyped with 14 microsatellite loci with a subset of 211 fish being genotyped by mtDNA PCR-RFLP. In both catchments, populations were found to be weakly differentiated genetically, and were most consistent with the meta-population theory of evolution. Similarly, individual spatial autocorrelation analysis indicated that each major tributary within the catchments could be considered as a distinct management or conservation unit. In the Tweed dataset, however, limitations in the sample coverage across the catchment reduced the robustness of some findings. Historical stocking of the river Dart with fish from Scotland and Iceland is well-documented. The long-term implications of these activities on contemporary Dart populations were assessed by genotyping 177 fish from the donor populations using scale samples taken in the 1960s and comparing them to contemporary Dart populations by undertaking admixture analysis. Overall, admixture between the donor and recipient populations was low and appeared to reflect natural underlying levels of genetic relationships. However, increased admixture of donor stocks with one extant Dart population was apparent, indicating some potentially long-term localised success of the stocked fish through hybridisation with the native populations; nevertheless, with the population continuing to decline, this should not be viewed as a successful supplementation programme. Two tributaries on the river Tweed, the Gala and Leader, were inaccessible to salmon for long periods due to the construction of barriers to migration. On both tributaries, fish passes were installed in the 1940s and re-colonisation of the tributaries was possible. Assignment analysis was undertaken and indicated that, contrary to findings for between catchment studies, salmon straying from the most proximate tributaries (i.e. the Ettrick and Caddon) did not appear to be the principal colonisers of the current Gala and Leader populations. Rather, the highest proportion of Gala samples assigned to the Teviot (42%), with the Leader populations assigning to many tributaries across the catchment (Ettrick 28%; Upper 21%; Teviot 19%). However, given the relatively weak differentiation of the baseline samples and limitations inherent in the dataset, the correct self-assignment of baseline samples was very low (average 26%; range 0-47%), hence interpretation must be undertaken with caution. Nevertheless, the findings suggest that the Gala population may have reached a temporally stable state in the 60 years since it has been accessible to salmon. Whilst the relatively small scale of these studies is acknowledged, the application of the findings in management and conservation of the species are discussed in a wider context. These studies would support the following recommendations: to include information on the historic (refugial) origin of contemporary populations in regional management strategies; to treat each major tributary as a distinct unit as an appropriate scale for catchment level management; and, with stocking and supplementation programmes appearing to have no significant long-term success, coupled with the relative speed with which extirpated tributaries appear to be naturally re-colonised, the use of stocking and supplementation programmes should be discouraged.
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Liu, Sixin. "Molecular marker analysis of adult plant resistance to powdery mildew in common wheat." Diss., Virginia Tech, 1999. http://hdl.handle.net/10919/11236.
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Powdery mildew, caused by Blumeria graminis (DC.) E.O. Speer f. sp. tritici E'm. Marchal (syn. Erysiphe graminis f. sp. tritici), is one of the major diseases of wheat (Triticum aestivum L.) worldwide. The use of cultivars with resistance to powdery mildew is an efficient, economical and environmentally safe way to control powdery mildew. Race-specific resistance has been extensively used in breeding programs; however, it is ephemeral. Adult plant resistance (APR) to powdery mildew is more durable as demonstrated by the cultivar Massey, which has maintained its APR to powdery mildew since its release in 1981. To develop an efficient breeding strategy, it is essential to understand the genetic basis of APR. The objectives of this study were to identify molecular markers associated with APR to powdery mildew in common wheat Massey and to verify their association using recombinant inbred (RI) lines.
A cross was made between the powdery mildew susceptible cultivar Becker and Massey. One hundred and eighty F2:3 lines were rated for disease severity under natural pressure of powdery mildew in field. Using both restriction fragment length polymorphism (RFLP) and microsatellite markers, three quantitative trait loci (QTL), designated as QPm.vt-1B, QPm.vt-2A and QPm.vt-2B, were identified in the Becker x Massey F2:3 generation. These loci are located on chromosomes 1B, 2A and 2B, respectively, and explained 17%, 29% and 11% of the total variation among F2:3 lines for powdery mildew resistance, respectively. Cumulatively, the three QTLs explained 50% of the phenotypic variation among F2:3 lines in a multi-QTL model. The three QTLs associated with APR to powdery mildew were derived from Massey and displayed additive gene action. QPm.vt-2B also fits a recessive model for APR to powdery mildew.
In the second part of this study, 97 RI lines were developed from the Becker x Massey cross. The RI lines were evaluated for APR to powdery mildew under natural disease pressure for three years. Both single marker analysis and interval mapping confirmed the presence of the three QTLs identified in the F2:3 generation. The three QTLs, QPm.vt-1B, QPm.vt-2A and QPm.vt-2B, accounted for 15%, 26% and 15% of the variation of mean powdery mildew severity of the RI lines over three years. In a multi-QTL model, the three QTLs explained 44% of the phenotypic variation of the RI lines. The RI lines were grouped according to the genotype of the three QTLs, represented by markers GWM304a, KSUD22 and PSP3100, respectively. The RI lines with Massey alleles at all three loci had a mean disease severity of 3.4%, whereas the RI lines with Becker alleles at all three loci had a mean disease severity of 22.3%. These severity values are similar to those of the corresponding parents.
The molecular markers identified and verified as to their association with APR to powdery mildew in Massey have the potential for use in marker-assisted selection for resistance to powdery mildew and in pyramiding powdery mildew resistance genes, as well as facilitating a better understanding of the molecular basis of APR to powdery mildew.Ph. D.
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Villanueva, Jorge Luis Chacón. "Epidemiologia molecular do vírus da laringotraqueíte infecciosa isolados de surtos em poedeiras comerciais no Estado de São Paulo." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/10/10133/tde-23012009-155035/.
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Este estudo teve como objetivo detectar e caracterizar molecularmente isolados de campo do vírus da laringotraqueíte infecciosa (VLTI) detectados de aves comerciais com e sem sintomatologia da doença de diferentes regiões do Estado de São Paulo. O estudo incluiu amostras coletadas durante e após o primeiro surto epidêmico da laringotraqueíte infecciosa (LTI) no Brasil, região de Bastos, e de outras localidades durante o período de 2002-2008. A caracterização molecular foi realizada através da técnica de polimorfismo do comprimento de fragmentos de restrição (RFLP) de produtos da reação em cadeia pela polimerase (PCR) dos genes da glicoproteína E, G, proteína quinase (TK) e da proteína reguladora da transcrição (ICP4), assim como pela análise de seqüências dos genes TK e ICP4. Para seqüenciamento do gene ICP4 foram desenvolvidas duas reações de PCR que amplificam dois diferentes fragmentos do gene ICP4. As técnicas de PCR-RFLP e seqüenciamento de DNA mostraram resultados idênticos, diferenciando os isolados de campo das cepas vacinais de origem de cultivo celular (TCO) e de embrião de galinha (CEO). Os resultados mostraram que o surto clínico na região de Bastos foi causado por uma cepa não vacinal e de alta virulência (cepa Bastos), embora também tenha sido possível detectar dois isolados relacionados à cepa CEO circulando durante o surto. Verificou-se que a cepa causadora do surto severo na região de Bastos continua circulando na região apesar do uso de vacinas atenuadas. Além disso, foram isoladas amostras relacionadas às cepas CEO e à cepa Bastos apartir de granjas comerciais de poedeiras localizadas fora da região de Bastos e que estão envolvidas em quadros clínicos da doença. Este estudo mostra (1): a persistência do vírus selvagem de campo na região de Bastos (cepa Bastos) apesar das medidas de controle e do uso de vacinas atenuadas, (2): a disseminação da cepa Bastos e das cepas vacinais CEO para outras regiões, e (3): a eficacia da estratégia padronizada neste estudo para a caracterização e diferenciação de isolados de campo e de cepas vacinais.The objective of this study was the molecular detection and characterization of field isolates of infectious laryngotracheitis virus (ILTV) detected from commercial chickens with and without clinical signs of the disease from regions of the São Paulo state. The study included samples collected during and after of the first epidemic infectious laryngotracheitis (ILT) outbreak in Brazil, Bastos region, and from other regions during 2002-2008. The molecular characterization was developed by restriction fragment length polymorphic analysis (RFLP) of polymerase chain reaction (PCR) products of glycoprotein E, G, thymidine kinase (TK) and regulatory protein of transcription (ICP4) gene, and by sequence analysis of TK and ICP4 gene. For ICP4 gene sequencing, two PCR assays have been developed for amplification of two different fragments of ICP4 gene. The PCR-RFLP and DNA sequencing techniques showed identical results, they could differentiate the field isolates from vaccine strains, tissue culture origin (TCO) and chicken embryo origin (CEO). The results showed that the severe outbreak in Bastos region was caused by a non-vaccine and virulent strain (Bastos strain); however it was possible to detect two isolates closely related to the CEO vaccine strain circulating during the outbreak. This study showed that the strain, which it caused the severe outbreak in Bastos region continue circulating in these region despite of the use of attenuate vaccines. In addition, the present research showed that isolates related to CEO and Bastos strains are circulating in commercial layer flocks located outside the Bastos region, and were involving in clinical cases of the disease. This study shows (1) the persistence of the wild field strain in Bastos region (Bastos strain) despite of the control measures and the use of attenuate vaccines, (2) the dissemination of the Bastos and CEO strains to other regions, and (3) the efficacy of the strategy standardized in this study to characterization and differentiation of field isolates and vaccine strain.
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Gustafson, Aubree Marie. "T-RFLP analysis of bacterial 16S rRNA gene sequences isolated from river otter (Lontra canadensis) scat and parasite screening for the presence of Toxoplasma gondii." Scholarly Commons, 2009. https://scholarlycommons.pacific.edu/uop_etds/732.
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In order to analyze the bacterial community of river otter scat (fecal material) at the class level, river otter scat samples were collected from Grizzly Island Wildlife Area (Solano County, CA) and the Cosumnes River Preserve (Sacramento County, CA). DNA was isolated from each sample with the MOBIO PowerSoil™ DNA Isolation Kit and 16S rRNA gene sequences were amplified from each sample. After digestion with Mspl, TRFLPs were analyzed in an ABI Prism™ 310 Genetic Analyzetin triplicate and data peak information from each electropherogram was uploaded into the Phylogenetic Assignment Tool (PAT). Species belonging to the Class Bacilli were the most abundant followed by unclassified species.
Two road killed river otters were necropsied to recover brain and blood tissue. DNA was isolated using the Qiagen Tissue DNeasy Kit. Samples from both otters were amplified with a singe tube nested PCR primer set for the detection of the ITS 1 region of Toxoplasma gondii. Scat samples used in the T-RFLP analysis were also tested for the presence ofT. gondii using the same nested primer set. Neither the river otter tissue samples nor any of the scat samples used in this analysis showed evidence of infeGtion with T. gondii.
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Siafakas, Nikolaos. "Molecular classification of coxsackie A virus reference and wild type strains on the basis of RFLP analysis and sequencing of the 5'-UTR : structural and evolutionary aspects." Thesis, University of Essex, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.369367.
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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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Baldwin, Quentin F. "Effects of prescribed burning upon mycorrhizal fungal diversity inhabiting the roots of two and a half-year old black spruce (Picea mariana) : molecular characterization of ectomycorrhizal fungi via PCR/RFLP analysis /." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ42348.pdf.
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Palíková, Petra. "Kvasinky a víno." Master's thesis, Vysoké učení technické v Brně. Fakulta chemická, 2010. http://www.nusl.cz/ntk/nusl-216597.
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This thesis deals with isolation and identification wine yeasts from grapes and must. For analysis was used white wine Sauvignon that was grown and producing after needs ecological agriculture. Remove samples were processed in laboratory and by the help of dilution method were obtained pure culture isolated yeasts. In the following step, by the application of commercial kit UltraCleanTM Microbial DNA Isolation Kit we were able to isolated individual DNA that it was used to the next analysis. Isolated DNA was amplification by PCR method with ITS1 and ITS4 primers. PCR products were detected on agarose gel. Amplification samples were chopped five restriction endonucleases: HaeIII, HinfI, TaqaI, AluI and MseI. Chopped DNA was detected by the same way as PCR products and it was compared with restriction patterns of collection yeasts. In the next step it was compared genetic similarity of isolated yeasts by using BioNumerics software. As a criterion it was used Pearson coefficients and UPGMA clastering analysis. The result is dedrogram of genetics similarity isolated yeasts.
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松田, 陽介, and Yosuke MATSUDA. "モミ根系における外生菌根菌の群集生態学的研究." 名古屋大学農学部付属演習林, 1999. http://hdl.handle.net/2237/8572.
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McTavish, Sharla. "Comparative analysis of New Zealand campylobacter isolates using MLST, PFGE and flaA PCR RFLP genotyping : a thesis submitted to the Victoria University of Wellington in partial fulfilment of the requirements for the degree of Master of Science in Molecular Microbiology /." ResearchArchive@Victoria e-Thesis, 2008. http://hdl.handle.net/10063/413.
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Cavell, Jane Sarah. "Cytogenetic and RFLP analyses of somaclonal variation in Nicotiana tabacum." Thesis, University of Bath, 1989. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.328517.
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Schmidt, Maren [Verfasser]. "Die Aussagekraft zusätzlicher RFLP-Analysen in defizienten Abstammungsbegutachtungen mittels Short-Tandem-Repeat-Analyse / Maren Schmidt." Kiel : Universitätsbibliothek Kiel, 2012. http://d-nb.info/1020283599/34.
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Moreira, Abdiel Aparecido. ""Pesquisa de sítios de restrição enzimática em segmento da ORF K1 do genoma de herpesvírus humano tipo 8 (HHV-8) em isolados clínicos de São Paulo: relação com subtipos virais e implantação da técnica de RFLP (Restriction Fragment Length Polymorphism Analyses) para determinar subtipos virais"." Universidade de São Paulo, 2003. http://www.teses.usp.br/teses/disponiveis/9/9136/tde-01102004-164856/.
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A epidemia da Síndrome de Imunodeficiência Adquirida (AIDS) fez aumentar a incidência de sarcoma de Kaposi (SK) em todos os países e o SK passou a ser considerado doença definidora de AIDS. Desde a descoberta de seu agente etiológico, o herpesvírus humano tipo 8 (HHV-8), vários estudos vêm sendo realizados com o objetivo de caracterizar subtipos virais presentes em todas as formas de SK: clássica, endêmica, iatrogênica e epidêmica. Os sistemas rotineiramente usados na subtipagem do HHV-8 têm usado o seqüenciamento de um gene que contém regiões hipervariáveis e que codifica uma glicoproteína de membrana viral (ORF K1). O presente trabalho apresenta um sistema de subtipagem alternativo que se baseia na presença de sítios de restrição enzimática em um pequeno segmento do gene ORF K1, região hipervariável 1 (VR1) e que permite discriminar subtipos virais. A análise de 68 seqüências; 50 que pertenciam a 36 pacientes com sarcoma de Kaposi infectados e não infectados pelo HIV-1 de São Paulo e 18 a protótipos dos subtipos A a E, mostraram mapas de restrição enzimática característicos dos principais subtipos virais descritos até o momento. Tomando como base apenas as enzimas disponíveis comercialmente, foram selecionadas cinco que se mostraram úteis para a subtipagem de HHV-8: Taq I, Nsi I, Hinf I, Hae III e Mse I. Os resultados obtidos com a técnica de PCR-RFLP (reação em cadeia de polimerase associada à análise do polimorfismo de fragmentos de restrição enzimática) mostraram que de 48 espécimes brasileiros previamente classificados como sendo dos subtipos A, B e C por seqüenciamento gênico, todos foram corretamente subtipados pela técnica de PCR-RFLP. Três amostras (duas do subtipo A e uma do B) apresentaram mais um sítio de restrição enzimática além dos descritos como sendo os predominantes. Mais recentemente, outras 27 amostras de 18 casos de infecção por HHV-8 foram subtipadas pela PCR-RFLP. Houve detecção de oito isolados do subtipo A, sendo seis de variante predominante, um de variante minoritária conhecida e um de nova variante viral. Dois casos de infecção por HHV-8 do subtipo B e sete do subtipo C também foram identificados. Finalmente, um provável caso de infecção pelo subtipo E foi encontrado em paciente com SK-AIDS disseminado, resistente à quimioterapia. Na avaliação global, houve maior número de casos de infecção por HHV-8 dos subtipos A e C. Concluindo, devido à alta sensibilidade e especificidade, baixo custo, rapidez e facilidade de execução, a técnica de PCR-RFLP pode ser usada em larga escala para estudos de epidemiologia molecular, principalmente em países em desenvolvimento.AIDS epidemic has increased the incidence rates of Kaposi s sarcoma (KS) in all countries, and KS has been considered an AIDS-defining illness. Since the discovery of the human herpesvirus 8 (HHV-8), the etiological agent of KS, several studies have been conducted in order to characterize HHV-8 in all forms of KS: classic, endemic, iatrogenic, and epidemic. The HHV-8 genome presents a hypervariable region termed ORF K1 useful for virus subtyping. The objectives of the present study were to describe an alternative method for subtyping HHV-8, to compare this new method with DNA sequencing, and to use this method for HHV-8 subtyping. After cloning and sequencing a segment of the ORF K1 (VR1) in 50 HHV-8/DNA isolates from 36 Brazilian KS-AIDS patients, we searched for restriction enzymatic sites in this segment of DNA, and compared them with 18 sequences reported in the literature. Then we constructed the enzymatic restriction maps useful for discriminating all HHV-8 subtypes described up to now, and standardized a PCR-RFLP (restriction fragment length polymorphism analysis) using five commercial enzymes: Taq I, Nsi I, Hinf I, Hae III e Mse I. After comparing the results obtained by the two methods, we used PCR-RFLP for HHV-8 subtyping in 27 new HHV-8/DNA isolates. The results obtained by DNA sequencing and PCR-RFLP showed 100% of concordance, and allowed the use of PCR-RFLP for HHV-8 subtyping. Indeed, we disclosed that among KS-AIDS patients from São Paulo, subtypes A and C are more prevalent than subtype B. Although additional restriction sites were detected in some Brazilian HHV-8 isolates, the majority of them belonged to the predominant strains described in the literature. Interestingly, one probable case of HHV-8 subtype E was detected in a patient who presented disseminated KS and resistance to chemotherapy. Because of its high sensitivity, specificity, low cost, and rapid execution, PCR-RFLP could be used on a large scale, mostly in countries with poor resources and where KS is endemic.
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Benitez, Maria Soledad. "Applied T-RFLP Analyses for the Identification and Characterization of Microbial Populations Associated With Damping-Off Incidence in a Transitional Organic Cropping System." The Ohio State University, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=osu1218471106.
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Kirker, Grant Terral. "EFFECTS OF CHLOROTHALONIL AND BUTYLATED HYDROXYTOLUENE ON MICROBIAL COMMUNITIES INVOLVED IN THE DETERIORATION OF WOOD USING TERMINAL RESTRICTION FRAGMENT LENGTH POLYMORPHISM (T-RFLP) ANALYSES." MSSTATE, 2008. http://sun.library.msstate.edu/ETD-db/theses/available/etd-04022008-155301/.
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The effects of an organic biocide (CTN) with and without co-added antioxidant (BHT) on microbial communities in SYP were assessed using terminal restriction fragment length polymorphism (T-RFLP) analyses in both field and accelerated decay laboratory studies. Ammoniacal copper quaternary (ACQ-C) was used as a positive control in the field study component, but not in the laboratory test. Field stakes were treated with 0.25 and 0.37% ammoniacal copper quat (ACQ-C), CTN (0.1 and 0.25%), CTN (0.1 and 0.25%) with 2% BHT added, 2% BHT alone, and controls were left untreated. In the field studies, preservative treatment slowed the initial colonization of wood by fungi. Higher species richness and diversity were found in non-biocidal treatments (BHT and untreated controls). Fungal communities in treated wood were different based on their species composition, but eventually became more similar to untreated controls. Preservative treatment increased richness and diversity of basidiomycete fungi, but overall presence of basidiomycetes was low compared to other fungi. Preservatives did not change the species composition of basidiomycetes compared to untreated controls. Preservative treatment initially increased bacterial richness and diversity, but over time these trends diminished to levels consistent with untreated controls. Preservatives changed the species composition of colonizing bacteria so that treated and untreated communities remained different over 15 months of soil exposure. Bacterial diversity was negatively correlated with CTN depletion at the lowest rate. In the accelerated decay laboratory test, the effects of CTN and/or BHT on bacterial, fungal, and basidiomycete communities in composted and uncomposted soil were evaluated over a 12 month period. Composted soil had less fluctuation in changing microbial diversity due to more constant moisture. The consensus of the analyses of the bacterial, fungal, and basidiomycete communities indicate that wood preservatives increased microbial species richness and diversity. Preservative treatment increased species turnover that decreased over time. Eventually, microbial communities approached a stable community structure consistent with untreated controls. Preservatives were completely degraded after 30 days exposure; however, definite changes in bacterial and fungal richness, diversity, and species composition were found. Basidiomycetes again represented the smallest portion of the microbial community involved in the overall decay process.
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Masoudi, Mehrnoush. "Identification of variants within the coding region and 5'-flanking region of the k-casein encoding gene in Holsteins using PCR-RFLP and PCR-SSCP analyses." Thesis, McGill University, 1996. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=23409.
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Single-strand conformation polymorphism analysis (SSCP) and restriction fragment length polymorphism analysis (RFLP) were used to determine the genotype of Holsteins at the $ kappa$-casein ($ kappa$-CN) locus. A 432-bp fragment within exon IV containing nucleotide substitutions diagnostic of the A- and B-variants of $ kappa$-CN was amplified using the polymerase chain reaction (PCR). The sires from the earliest years of the AI industry had a significantly higher (p $<$ 0.01) frequency of allele than sires in modern usage. These data indicate that selection or milk production parameters may discriminate against the B-allele. SSCP analysis was also used for detecting polymorphisms within the regulatory region of $ kappa$-CN gene. A 640-bp fragment within the 5$ sp prime$-flanking region of bovine $ kappa$-CN gene which contained the TATA box, CAAT box, and exon I was amplified using PCR. The SSCP analysis of this fragment revealed no variation, possibly due to the lower detection efficiency of SSCP with large fragment size. Nested primers were, therefore, designed to amplify fragments of 234- and 486-bp. Polymorphism was detected only in the 486-bp fragment and the two variants were designated M$ sb1$ and M$ sb2.$ The allelic frequencies of M$ sb1$ and M$ sb2$ in bulls used by AI industry before 1970 were 0.67 and 0.33, and in bulls used by AI industry after 1980 the frequencies were 0.68 and 0.32, respectively. The frequency of these alleles were not significantly different in Holsteins used by AI industry before 1970 and after 1980. Unlike the apparent change in frequency of the A- and B-variants noted within exon IV, this polymorphism seems to have not responded to selection. However, a higher frequency of M$ sb1$ allele appeared to be associated with B-variant (exon IV) genotypes. The presence of these variants within the regulatory region may possibly be involved in the quantitative expression of $ kappa$-CN gene. (Abstract shortened by UMI.)
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Kirker, Grant Terral. "Effects of chlorothalonil (CTN) and butylated hydroxy-toluene (BHT) on microbial communities involved in the deterioration of wood using terminal restriction fragment length polymorphism (T-RFLP) analyses." Diss., Mississippi State : Mississippi State University, 2008. http://library.msstate.edu/etd/show.asp?etd=etd-04022008-155301.
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Bleul, Catrin. "Molekularbiologische Analyse mikrobieller Gemeinschaften in Talsperrensedimenten." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2004. http://nbn-resolving.de/urn:nbn:de:swb:14-1097570982718-83940.
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Mikrobielle Prozesse spielen eine wichtige Rolle im Sediment von Talsperren und Seen. Demgegenüber stehen nur unzureichende Erkenntnisse über die Zusammensetzung mikrobieller Biozönosen in Sedimenten sowie deren Aktivität zur Verfügung. Das Ziel dieser Studie war die Untersuchung und der Vergleich der Zusammensetzung und der Struktur mikrobieller Gemeinschaften in Sedimenten um eine Abschätzung der mikrobiellen Diversität in Talsperrensedimenten unterschiedlicher Trophie zu erreichen. Durch die Kombination der in dieser Arbeit verwendeten Methoden (Vergleichende 16S rDNA Analyse, Fingerprinttechniken, klassische Methoden) konnte eine Charakterisierung der mikrobiellen Zusammensetzung der obersten 5 cm von den Talsperrensedimenten Neunzehnhain, Muldenberg, Quitzdorf und Saidenbach erzielt werden. Die vergleichende 16S rDNA Analyse offenbarte in 2541 analysierten rekombinanten Klonen 528 verschiedene Sequenztypen, welche zu 293 OTUs zusammengefaßt werden konnten. Obwohl die Gemeinschaften der verschiedenen Talsperren nur schwach auf der Ebene der phylogenetischen Gruppen differierten, konnte durch die Verwendung von Ähnlichkeitsindices gezeigt werden, dass jede Talsperre eine spezifische mikrobielle Sedimentgemeinschaft aufweist. Über 60% aller Klone zeigten Ähnlichkeiten von mehr als 97% zu 16S rDNA-Sequenzen kultivierter Organismen oder phylogenetisch eingeordneten Sequenzen (14 bekannte phylogenetische Gruppen). Alle anderen Klone zeigten hohe Sequenzhomologien zu unidentifizierten, phylogenetisch bisher nicht eingeordneten Bakterien. Diese Bakterien waren mit Anteilen zwischen 19,8% (Muldenberg) und 54,6% (Saidenbach) in den 16S rDNA Bibliotheken repräsentiert. Mittels Fingerprinttechniken (DGGE, T-RFLP, ARISA) konnten komplexe Muster der mikrobiellen Diversität erzeugt werden. Dabei konnten die Ergebnisse der 16S rDNA Analyse bestätigt werden. Durch die verwendeten Methoden konnte eine komplexe mikrobielle Diversität in den Sedimenten aufgedeckt werden und die Ergebnisse weisen darauf hin, dass die mikrobielle Diversität in Sedimenten wesentlich höher ist als bisher angenommen.
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Eschenhagen, Martin. "Molekulare Untersuchung zweier Belebtschlammanlagen unter besonderer Berücksichtigung der biologischen Phosphorelimination." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2004. http://nbn-resolving.de/urn:nbn:de:swb:14-1091188675328-95596.
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Aufgrund der ökologischen und ökonomischen Problematik der chemischen Phosphatfällung ist eine Optimierung der Effizienz und Stabilität der biologischen Verfahren zur Phosphat-elimination erforderlich. Hierfür ist jedoch ein fundiertes Wissen über die daran beteiligten Organismen eine entscheidende Vorraussetzung. Das Ziel der vorliegenden Arbeit war es, die mikrobielle Populationstruktur von zwei Belebtschlamm-anlagen im Labormaßstab mit Hilfe von drei unterschiedlichen 16S rDNA basierenden molekular-biologischen Methoden zu charakterisieren. Ein besonderer Schwer-punkt ist hierbei die Analyse der Bakterien, die mit der erhöhten biologischen Phosphat-elimination in Verbindung gebracht werden. Dies sind Vertreter der Rhodocyclus-Gruppe, der Gattung Tetrasphaera und der Gattung Acinetobacter. Als Untersuchungsobjekte wurden zwei Hauptstromverfahren zur erhöhten biologischen Phosphatelimination gewählt, die sich im Schlamm-alter, der Schlammbelastung und der sich daraus resultierenden Nitrifikationsleistung unterscheiden. Aufgrund der gewählten Verfahrensweisen wurde der Einfluss der Nitrifikation auf die Zusammensetzung der Belebtschlammbiozönose ebenfalls untersucht. Um praxisnahe Verhältnisse zu erreichen, wurden die Anlagen mit kommunalem Abwasser beschickt. Für einen Vergleich sollten Proben aus kommunalen Kläranlagen mit deutlich anderen Verfahrensweisen in die Untersuchungen mit einbezogen werden.
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Schleicher, Anna Lara Verfasser], and Eckhard [Akademischer Betreuer] [Wolf. "Analysis of OCT4 expression in transgenic porcine embryos carrying an OCT4-RFP reporter construct / Anna Lara Schleicher ; Betreuer: Eckhard Wolf." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2017. http://d-nb.info/1137226927/34.
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Schleicher, Anna Lara [Verfasser], and Eckhard [Akademischer Betreuer] Wolf. "Analysis of OCT4 expression in transgenic porcine embryos carrying an OCT4-RFP reporter construct / Anna Lara Schleicher ; Betreuer: Eckhard Wolf." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2017. http://d-nb.info/1137226927/34.
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Eschenhagen, Martin. "Molekulare Untersuchung zweier Belebtschlammanlagen unter besonderer Berücksichtigung der biologischen Phosphorelimination." Doctoral thesis, Technische Universität Dresden, 2003. https://tud.qucosa.de/id/qucosa%3A24360.
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Aufgrund der ökologischen und ökonomischen Problematik der chemischen Phosphatfällung ist eine Optimierung der Effizienz und Stabilität der biologischen Verfahren zur Phosphat-elimination erforderlich. Hierfür ist jedoch ein fundiertes Wissen über die daran beteiligten Organismen eine entscheidende Vorraussetzung. Das Ziel der vorliegenden Arbeit war es, die mikrobielle Populationstruktur von zwei Belebtschlamm-anlagen im Labormaßstab mit Hilfe von drei unterschiedlichen 16S rDNA basierenden molekular-biologischen Methoden zu charakterisieren. Ein besonderer Schwer-punkt ist hierbei die Analyse der Bakterien, die mit der erhöhten biologischen Phosphat-elimination in Verbindung gebracht werden. Dies sind Vertreter der Rhodocyclus-Gruppe, der Gattung Tetrasphaera und der Gattung Acinetobacter. Als Untersuchungsobjekte wurden zwei Hauptstromverfahren zur erhöhten biologischen Phosphatelimination gewählt, die sich im Schlamm-alter, der Schlammbelastung und der sich daraus resultierenden Nitrifikationsleistung unterscheiden. Aufgrund der gewählten Verfahrensweisen wurde der Einfluss der Nitrifikation auf die Zusammensetzung der Belebtschlammbiozönose ebenfalls untersucht. Um praxisnahe Verhältnisse zu erreichen, wurden die Anlagen mit kommunalem Abwasser beschickt. Für einen Vergleich sollten Proben aus kommunalen Kläranlagen mit deutlich anderen Verfahrensweisen in die Untersuchungen mit einbezogen werden.
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Nguyen, Thi Diem Hong. "Etude par Raman-Laser-Fibres Optiques (RFLO) de la composition chimique des calculs urinaires en vue de son application au domaine biomédical." Châtenay-Malabry, Ecole centrale de Paris, 1993. http://www.theses.fr/1993ECAP0292.
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La méthode Raman-Laser-Fibres Optiques (RLFO) a été utilisée pour étudier la possibilité d'une analyse in vivo des calculs urinaires. Cette dernière est indispensable pour guider les ondes de chocs dans la fragmentation des calculs par voie endoscopique ou extracorporelle ainsi que pour mieux définir les traitements préventifs contre les récidives lithiasiques. Nous avons d'abord crée sur ordinateur, une bibliothèque des spectres Raman des 48 constituants lithiasiques et de 330 de leurs mélanges binaires et ternaires les plus fréquents, ce qui nous a permis d'effectuer des analyses semi-quantitatives des calculs à partir de leurs spectres RLFO. Pour améliorer la qualité des spectres RLFO, une optode à filtres a été mise au point. Cette optode permet de mettre en évidence l'effet de filtres optiques dans l'élimination des signaux parasites provenant des fibres optiques. Nous avons également crée puis testé des modelés d'analyse quantitative par la méthode PCR (Principal Component Regression) de mélanges synthétiques de constituants lithiasiques (mélanges binaires struvite-carbonate apatite et mélanges ternaires acide urique anhydre-urate acide de sodium anhydrewhewellite) pour les spectres Raman directs et pour les spectres RLFO. Les résultats sont satisfaisants pour une exploitation clinique de l'analyse des calculs urinaires: selon les modèles, l'erreur quadratique moyenne de prédiction pour des mélanges inconnus se situe entre 1,5 et 2,6%. Enfin, l'étude de la répartition des constituants au sein d'un calcul hétérogène est également rendue possible par la technique de cartographie RLFO. De tels renseignements permettraient de déterminer les facteurs impliqués dans la genèse des calculs urinaires et constitueraient une aide précieuse pour les médecins dans l'étude étiologique de la maladie lithiasique.
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Lelièvre, Stéphanie. "Identification et caractérisation des frayères hivernales en Manche Orientale et la partie sud de la mer du Nord : Identification des oeufs de poissons, cartographie et modélisation des habitats de ponte." Nantes, 2010. http://www.theses.fr/2010NANT2110.
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Une meilleure connaissance des frayères des principaux poissons commerciaux de la mer du Nord semble nécessaire pour leur surveillance. La composition et l'abondance des espèces d'oeufs collectés par le CUFES (Continuous Underway Fish Egg Sampler) sont comparées à celle collectées par le VET (Vertical Egg Trawl) permettant de prouver l'efficacité du CUFES en Manche et mer du Nord. L'identification des œufs de poissons principalement basée sur des critères morphologiques n'est pas toujours fiable. En effet, certaines espèces comme la morue (Gadus morhua) et le merlan (Merlangius merlangus) ont la même gamme de taille, ainsi des méthodes altenatives ont été développées. Premièrement, une technique de biologie moléculaire par PCR-RFLP puis un nouveau système d'analyse d'images, le ZooScan ont été développés pour identifier les œufs de poissons. Des cartes annuelles des frayères hivernales ont été réalisées et comparées entre elles afin de déterminer des zones de ponte récurrentes. Les œufs sont généralement bien distribués sur la zone d'étude, à l'exception de la zone Nord-Ouest de la mer du Nord, près des côtes écossaises. Et enfin, l'habitat de ponte des poissons a été modélisé en utilisant les méthodes GLM (Generalised Linear Model) et RQ (Regression Quantile) en fonction des paramètres environnementaux disponibles afin de prédire les frayères. Les résultats de cette étude multidisciplinaire ont permis d'améliorer les connaissances sur les frayères hivernales en Manche Orientale et sud mer du Nord et ont été discutés dans une perspective de protection et de conservation de ces zonesA better knowledge and monitoring of principal commercial fish spawning grounds have become necessary in the North Sea. The efficiency of CUFES was proved by sampling pelagic fish eggs in winter in Eastern Channel and Southern North Sea. Fish egg taxonomic identification based on visual criteria cannot always be carried out effectively. In particular, cod (Gadus morhua), and whiting (Merlangius merlangus) or flounder (Platichthys flesus) and dab (Limanda limanda) have the same range of egg diameter and similar morphologies. Alternative identification methods using molecular techniques were developed to improve the accuracy of egg taxonomic identification. First, PCR-RFLP method, then, in order to accelerate egg identification, the use of a new laboratory imaging system, the ZooScan, able to produce high resolution images of zooplankton samples, was adapted to fish eggs and allower their automated identification using supervised learning algorithms. The location of winter spawning grounds of fishes in the Southern North Sea and the Eastern Channel was illustrated using yearly maps and analysed over the available period to define recurrent, occasional and unfavorable spawning areas. Generally, fish eggs were found over the study area, except for the North Western of the North Sea, near Scottish coasts. Important spawning areas were clearly localised along the Belgian, Dutch and Danish coasts. Habitat modelling of these fish spawning areas was carried out using both GLM (Generalised Linear Model) and QR (Regression Quantile) and associated egg abundance to physical conditions such as temperature, salinity, bedstress, chlorophyll a concentration and bottom sediment types to characterize spawning habitat conditions and predict their extent and location. The results of this approach improve the understanding of spawning grounds distribution and were discussed in the context of the protection and conservation of critical spawning grounds
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Bykov, Igor. "Experimental studies of materials migration in magnetic confinement fusion devices : Novel methods for measurement of macro particle migration, transport of atomic impurities and characterization of exposed surfaces." Doctoral thesis, KTH, Fusionsplasmafysik, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-145045.
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During several decades of research and development in the field of Magnetically Confined Fusion (MCF) the preferred selection of materials for Plasma Facing Components (PFC) has changed repeatedly. Without doubt, endurance of the first wall will decide research availability and lifespan of the first International Thermonuclear Research Reactor (ITER). Materials erosion, redeposition and mixing in the reactor are the critical processes responsible for modification of materials properties under plasma impact. This thesis presents several diagnostic techniques and their applications for studies of materials transport in fusion devices. The measurements were made at the EXTRAP T2R Reversed Field Pinch operated in Alfvén laboratory at KTH (Sweden), the TEXTOR tokamak, recently shut down at Forschungszentrum Jülich (Germany) and in the JET tokamak at CCFE (UK). The main outcomes of the work are: Development and application of a method for non-destructive capture and characterization of fast dust particles moving in the edge plasma of fusion devices, as well as particles generated upon laser-assisted cleaning of plasma exposed surfaces. Advancement of conventional broad beam and micro ion beam techniques to include measurement of tritium in the surfaces exposed in future D-T experiments. Adaption of the micro ion beam method for precision mapping of non uniform elements concentrations on irregular surfaces. Implementation of an isotopic marker to study the large scale materials migration in a tokamak and development of a method for fast non destructive sampling of the marker on surfaces of PFCs.QC 20140508
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Trape, Sébastien. "Les Mugilidae des côtes ouest-africaines : identification moléculaire, phylogénie et bio-écologie du recrutement des juvéniles dans un estuaire hypersalé." Montpellier 2, 2009. http://www.theses.fr/2009MON20163.
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La persistance de la sécheresse depuis les années 1970 en Afrique de l'Ouest a particulièrement affecté le fonctionnement des écosystèmes côtiers. Dans certains estuaires cette perturbation a engendré une inversion du gradient de salinité et l'apparition de zones hypersalées (salinité > 60). Afin d'appréhender l'impact de ces changements environnementaux sur le recrutement et la croissance des juvéniles dans les milieux estuariens, les patrons spatio-temporels de recrutement (structures de taille et distributions d'abondance) de toutes les espèces de mulets ont été étudiés dans l'estuaire inverse du Sine Saloum (Sénégal). La croissance des juvéniles a été mesurée grâce aux microstructures journalières des otolithes. La taxonomie des Mugilidae en Afrique de l'Ouest étant confuse et les critères morphométriques nécessaires à l'identification des juvéniles encore inconnus, nous avons dans un premier temps (1) élaboré une phylogénie moléculaire des espèces, (2) développé une méthode génétique pour l'identification des juvéniles et enfin (3) mis au point une clef d'identification morphométrique. Les analyses moléculaires ont révélé l'existence d'une nouvelle espèce sur les côtes ouest africaines (Chelon bandialensis) et montré que Mugil ashanteensis et Mugil metzeleaari précédemment confondues avec Mugil cephalus et Mugil curema constituent deux espèces valides. Les patrons spatio-temporels du recrutement des juvéniles ont révélé que Liza dumerili dominait très largement le peuplement en représentant 89 % des captures totales. Les décalages entre les périodes de recrutement des espèces expliquent la colonisation d'un même environnement par des espèces proches. La croissance des juvéniles de L. Dumerili, plus élevée avec les faibles salinités (moyenne 36), met en exergue les contraintes apportées par l'hypersalinisationThe persistence of the drought since the 1970s in West Africa has particularly affected the functioning of coastal ecosystems. This perturbation has conducted to reverse salinity gradients in some estuaries and the appearance of hypersaline areas (salinity > 60). To get some insights about the consequences of these environmental changes on the recruitment and the growth of juveniles in estuarine ecosystems, spatio-temporal patterns of recruitment (size structures and abundance distributions) of all Mugilidae species have been studied in the inverse Sine Saloum estuary (Senegal). The growth has been measured using otolith daily microincrements. The taxonomy of West African Mugilidae being confused and the morphological criteria necessary for identifying juveniles yet unknown, in a first step we have (1) carried out a molecular phylogeny of the mullet species, (2) developed a genetic method for identifying juveniles and finally (3) developed a morphometric identification key for juveniles. Molecular analyses have revealed the existence of a new mugilid species along the West African coasts (Chelon bandialensis) and shown that Mugil ashanteensis and Mugil metzeleaari previously confounded with Mugil cephalus and Mugil curema are valid species. The spatio-temporal patterns of recruitment of juveniles have revealed that Liza dumerili largely dominated the assemblage, representing 89% of the total catches. The gaps between recruitment periods of the species explain the colonisation of a same environment by close related species. The juvenile growth of L. Dumerili, higher in low salinities (mean 36), highlights constraints associated with hypersalinity
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Havlů, Monika. "Podnikatelský záměr rozvoje společnosti." Master's thesis, Vysoké učení technické v Brně. Fakulta podnikatelská, 2012. http://www.nusl.cz/ntk/nusl-223345.
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The present dissertation is the design of a company's business plan, with a view to the new monitoring system, with all the important factors such as economic and technical factors, and legislative changes in the branch are taken into consideration. Furthermore there is the view to the economic standing of the company is also evalua-ted. The business plan is simultaneously considered from a general view of potential implementation of the proposed solution.
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Liu, Win-Lin, and 劉文粦. "GISH, PCR-RFLP and RFLP analysis of intergeneric hybrids combining for Ascocenda and Phalaenopsis." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/25455442480587729275.
Full textAbstract:
碩士國立屏東科技大學
生物科技研究所
97
The Phalaenopsis is one of potential and high economic crop in Taiwan. The genetic inheritance of intergeneric hybrids combining Ascocenda John De Biase "Blue" (♀) and Phalaenopsis Chih Shang's Stripes (♂) (F1) were detected by genomic in situ hybridization (GISH), PCR-restriction fragment length polymorphism (PCR-RFLP) and restriction fragment length polymorphism (RFLP). The intergeneric hybrids (F1 plants) can be confirmed that from Ascocenda John De Biase "Blue" (♀) and Phalaenopsis Chih Shang's Stripes (♂) by GISH. No recombination can be found between two species. Twenty-seven randomly selected for following DNA anaysis. Both internal trandscribed spacer (ITS) DNA and external transcribed spacer (ETS) of nuclear ribosomal DNA (nrDNA) were separately analyzed by PCR-RFLP analysis. It is not in agreement between ITS and ETS analysis. In ITS analysis, those hybrids showed biparental patterns, however, they only showed the DNA pattern of maternal parent, Ascocenda John De Biase "Blue"(♀), in ETS analysis. Furthermore, the trnL intron of chloroplast genome (cpDNA) was also analysis by RCR-RFLP. The result shows that cpDNA is maternal inheritance in the intergeneric hybrids. For further realizing aforementioned incongruence of genetic inheritance between ITS and ETS by PCR-RFLP, RFLP analysis was used. The result shows that ETS region is also biparental inheritance same as that of ITS region. It indicates that the bias of PCR amplification for ETS region is happened in all hybids. After sequencing of ETS region for both parents, the bias of PCR amplification resulting of two-base mismatch between the primer designed in ETS region and the DNA template of Phalaenopsis Chih Shang's Stripes (♂) is suggested. In addition, seven hybrids show one unique DNA pattern of ETS by RFLP analysis. Inspection of the morphology of these seven hybrids, they show leaves than others. In final, GISH, PCR-RFLP and RFLP can be used for detecting the genetic inheritance of intergeneric hybrids. However, it should be kept from the primer competition in PCR amplification for mixed genome when we use PCR-RFLP analysis.
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Zhang, Jian Shi Ding, and 張簡士鼎. "RFLP analysis of deficiency on chromosome 8 in maize." Thesis, 1995. http://ndltd.ncl.edu.tw/handle/38172578209764465874.
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Lenz, Erin Jennifer. "Rhizobial T-RFLP analysis for differentiating soils and habitats." Diss., 2008. http://proquest.umi.com/pqdweb?did=1606926031&sid=2&Fmt=2&clientId=3552&RQT=309&VName=PQD.
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Chuang, Yueh-Feng, and 莊岳峰. "RFLP analysis on the genetic relationships of the wild soybeans in Taiwan." Thesis, 1999. http://ndltd.ncl.edu.tw/handle/29607804734545640193.
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碩士國立中興大學
農藝學系
87
The distribution of two soybean subgenus, Soja and Glycine, intersects on Taiwan. Therefore, a wide variety of wild soybean genetic resources can be found in Taiwan. This research is to understand the relationships between Taiwan wild soybean and its relatives. We used the total DNA of soybean plants as test material. Using two fragments of wild soybean''s mitochontrial DNA, atp6 and coxII, as probes, we performed RFLP analysis and, by the similarity coefficients among the accessions and the results of cluster analysis, investigated their genetic relationships. The result shows that, G. formosana, G. soja, G. gracilis, and G. max in subgenus Soja, the combinations of restriction enzyme－probe including BamHI－atp6, HindIII－coxII, and HindIII－atp6, all produced polymorphic bands. And we used the fragments of G. formosana and G. gracilis amplified by PCR reaction as probe to perform RFLP analysis. The larger than 2 kbp band amplified with the primer OPB-5 of G. gracilis as a probe, produced polymorphic DNA fragments; and the 600 bp band amplified with the primer OPB-12 of G. formosana as a probe, produced a specific band of G. formosana. By cluster analysis, we found that the genetic position of semi-wild G. gracilis lies in between G. max and G. soja, and that G. gracilis can be clustered with G. max. G. formosana lies in a group of its own due to the significant differences in DNA polymorphism and parentage between it and other species in subgenus Soja. In subgenus Glycine, only the DNA digested with BamHI using coxII as a probe to yield polymorphic hybridization production, which is also exclusive to G. dolichocarpa. The other combinations, including the 200 bp band amplified with the primer OPB-7 of G. dolichocarpa as a probe, produced no hybridization production. The results shows that there exists differences in DNA sequence among G. dolichocarpa, G. tomentella, and G. tabacina.
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SONG, SHU-LING, and 宋述玲. "Conservation at the 5'-region of human Phosphofructokinase gene by RFLP analysis." Thesis, 1990. http://ndltd.ncl.edu.tw/handle/23260812997164584935.
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Sulaiman, Yousuf Al-Wahaibi, and 尤瑟夫. "Detection of Salmonella choleraesuis in Taiwan Using PCR-RFLP and Antibiotics Resistant Analysis." Thesis, 2003. http://ndltd.ncl.edu.tw/handle/85555475913353558712.
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碩士國立屏東科技大學
熱帶農業暨國際合作研究所
91
Salmonella species is an important zoonotic pathogen in humans and animals that frequently caused worldwide food-borne gastrointestinal diseases transmitted from animals to human. Antimicrobial drug resistance was found to be increasing among S. choleraesuis strains in Pingtung, Taiwan. Using disc diffusion methods as recommended by the National Committee for Clinical Laboratory Standards (NCCLS) on Muellen—Hinton agar, tested antimicrobial susceptibility. Of the 101 isolates, 99% of the isolates showed resistance to carbenicillin, 97% to apramycin, 96% to ampicillin, and 96% to oxytetracyclin. Additionally, 87.1% of the isolates were resistant to gentamycin, 84.2% to neomycin, and 75.2% to chloramphenicol. However, 59.4% of the isolates were resistant to cefoperzone, and 8.9% to ceftiofur. We found also less than 5% of the isolates were resistant to norofloxacin, trimethoprim, and trimethoprim-sulfamethoxazol. Multiple resistances were observed among all of the isolates. All of the isolates tested showed resistance to at least three antibacterial drugs. Twenty-nine different drug resistance patterns were recorded. The most common patterns of antibiotic resistance (n= 39 isolates) were AmAprCfpCbCGmNT, followed by AmAprCbCGmNT (n=17). Using the PCR assay to determine the specificity of the primer iroB and invA in detecting S. choleraesuis at the genus level showed that all of 101 S. choleraesuis samples gave positive results. Only two of 101 S. choleraesuis samples gave positive results with integron specific primer pairs. However, comparing these two samples with other sample, the results showed that the difference laid in item resistance with other sample against the antimicrobials drugs of trimethoprim and trimethoprim-sulfamethoxazol. This study demonstrated that there was no relationship between the 29 different drug resistance patterns and the results of RFLP analysis of the PCR products using the primers of iroB and invA.
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Κράιτσεκ, Σπυριδούλα. "Γενετική ποικιλότητα και φυλογενετικές σχέσεις "λιμναίων" και "θαλάσσιων" πληθυσμών της Atherina boyeri." Thesis, 2006. http://nemertes.lis.upatras.gr/jspui/handle/10889/1112.
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Στην εργασία αυτή μελετήθηκε η γενετική δομή και οι φυλογενετικές σχέσεις μεταξύ έξι πληθυσμών της Atherina boyeri που προέρχονταν από τις περιοχές της Μυτιλήνης, της Νισύρου, της Κάσου, της Κύμης, και των λιμνών της Βιστωνίδας και Iznik (στην Τουρκία). Συγκεκριμένα, έγινε μελέτη των περιοριστικών θραυσμάτων ποικίλου μήκους (RFLP analysis) των τμημάτων 12S rRNA, 16S rRNA και του βρόγχου εκτόπισης (D-loop) του μιτοχονδριακού DNA. Τα αποτελέσματα αυτά συνδυάστηκαν με τα αποτελέσματα άλλων μελετών που αφορούσαν εννέα διαφορετικές περιοχές της Ελλάδας (Κάλυμνο, Κεφαλλονιά, Αμβρακικός, Κως, Λήμνος, Εύβοια, Ζάκυνθος, Λευκάδα και Κουρνά-Κρήτη) και είχαν γίνει στο εργαστήριο. Από την RFLP ανάλυση αποκαλύφθηκαν 23 διαφορετικοί σύνθετοι απλότυποι. Βάσει των αποτελεσμάτων γίνεται σαφής διαχωρισμός μεταξύ «λιμναίου» και «θαλάσσιου» τύπου πληθυσμών. Οι πληθυσμοί από τις λίμνες/λιμνοθάλασσες (Βιστωνίδα, Κουρνά, Κούταβος/Κεφαλλονιά, Αμβρακικός, Iznik/Τουρκία ), καθώς και από περιοχές που επικρατούν παρόμοιες συνθήκες (Κύμη, Βαθύ Καλύμνου), έχουν τους απλότυπους 1-6, ενώ οι «θαλάσσιου» τύπου πληθυσμοί (Κως, Λήμνος, Εύβοια, Μυτιλήνη, Νίσυρος, Κάσος, Ζάκυνθος, Λευκάδα) έχουν τους απλότυπους 7-23. Διαπιστώθηκε επίσης η ύπαρξη πέντε διαγνωστικών προτύπων μεταξύ «λιμναίων» και «θαλάσσιων» πληθυσμών. Επιπλέον, παρατηρήθηκε η ύπαρξη ενός διαγνωστικού προτύπου για τους πληθυσμούς από την Κω, τη Λήμνο και τη Νίσυρο, βάσει του οποίου μπορούμε να διαχωρίσουμε τους πληθυσμούς αυτούς από τους υπόλοιπους θαλάσσιους πληθυσμούς που μελετήθηκαν, καθώς και ένα διαγνωστικό πρότυπο βάσει του οποίου μπορούμε να διακρίνουμε τον πληθυσμό της Νισύρου από τους υπόλοιπους επτά πληθυσμούς «θαλάσσιου» τύπου. Με βάση τα δεδομένα αυτά υπολογίστηκε η καθαρή νουκλεοτιδική απόκλιση μεταξύ των πληθυσμών και βρέθηκε να είναι αρκετά υψηλή σε ορισμένες περιπτώσεις. Τα παραπάνω αποτελέσματα επιβεβαιώνονται και από τα δύο φυλογενετικά δένδρα που κατασκευάστηκαν με τις μεθόδους UPGMA και Μέγιστης Φειδωλότητας. Βάσει της τιμής του Nst (50%) που υπολογίστηκε μόνο η μισή από την ολική γενετική ποικιλότητα που παρατηρήθηκε οφείλεται σε διαφορές ανάμεσα στους πληθυσμούς, ενώ η υπόλοιπη οφείλεται σε ενδοπληθυσμιακές διαφορές.Τhe genetic differentiation and the phylogenetic relationships of six greek populations of Atherina boyeri were investigated at the mitochondrial level. The samples originated from the marine sites of Lesvos, Nisyros, Kasos, Kymi and the lakes Vistonida and Iznik (Turkey). RFLP analysis of three mtDNA segments (12S rRNA, 16S rRNA and D-loop) amplified by PCR were used. These results were combined with others available in the laboratory, concerning nine more greek populations (Kalymnos, Kefallonia, Amvrakikos, Kos, Limnos, Evvoia, Zakynthos, Leukada, Kourna/Crete). Twenty-three composite haplotypes where revealed from the RFLP analysis. There is a clear distinction between “marine” and “lagoon” type populations. In particular, the populations from the lakes/lagoons (Vistonida, Kourna, Kefallonia, Amvrakikos, Iznik), as well as the populations from sites with similar environmental conditions to the lakes/lagoons (Kymi, Kalymnos) have the haplotypes 1-6, while the “marine” type populations (Kos, Limnos, Evvoia, Lesvos, Nisyros, Kasos, Zakynthos, Leukada) have the haplotypes 7-23. Five specific restriction patterns were also revealed, which can be used to distinguish the “marine” from the “lagoon” type populations. Moreover, one diagnostic pattern, with which we can distinguish the populations from Kos, Limnos and Nisyros from the rest “marine” type populations studied, was revealed, as well as it was revealed one diagnostic pattern, with which we can distinguish the population of Nisyros from the rest “marine” type populations. The genetic divergence values estimated among “lagoon” and “marine” type populations were high, with the populations from Evvoia and Kourna to show the greatest divergence (10.450%) and the populations from Amvrakikos and Nisyros the lowest (5.549%). The above results were also confirmed and by the two phylogenetic trees that were conducted using the UPGMA and the Maximum Parsimony methods. The trees consist of two main clades, which contain the “marine” and “lagoon” populations respectively. Our results show that distinct “lagoon” populations (such as from Vistonida and Kourna) have similar genetic structure, a situation that is not true for the “marine” populations, since there are populations with completely different genetic structure. Finally, the Nst value (50%) indicates that half of the overall genetic diversity detected was between populations.
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Lashaway, Aubrey Rain. "Water quality and eukaryotic plankton dynamics in the Mission-Aransas Estuary, Texas from 2011-2012." 2013. http://hdl.handle.net/2152/22101.
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As the base of the food chain, plankton affect the cycling of nutrients and organic matter within ecosystems and support production at higher trophic levels. The overall goal of this project was to examine how natural water quality fluctuations, such as changes in nutrients, temperature, and salinity, influence estuarine plankton community structure. To achieve this, I examined water quality as well as the diversity and biomass of eukaryotic plankton communities in a subtropical estuary located within the Mission-Aransas National Estuarine Research Reserve. The sampling sites included in this study consisted of three bay (Copano Bay West, Copano Bay East, Aransas Bay) and two river (Mission River Estuary, Aransas River Estuary) estuary sites. Water samples were collected monthly at the five sites from September 2011 to August 2012 and analyzed for a suite of abiotic and biotic variables. Eukaryotic plankton diversity and community structure were evaluated by using the terminal restriction fragment length polymorphism (t-RFLP) method.
Although a narrow salinity gradient was present at the sampling sites, seasonal changes in water quality conditions were observed. In the river estuaries, water quality parameters defined three significant temporal periods at the Mission River Estuary site, whereas only one month differed at the Aransas River Estuary site, indicating little seasonal variation. The Copano Bay sites exhibited a seasonal pattern consisting of four periods, marked by a distinct fall (October, November, December) grouping, while Aransas Bay showed a seasonal pattern consisting of three periods, with no fall group. Even though the water quality conditions define different monthly groupings in the bay and river estuary sites, the same parameters – DOC, TDN, and pH – are the strongest drivers of the patterns at all of the sites.
Seasonal and spatial distinctions in the Mission-Aransas Estuary eukaryotic plankton community composition were determined using t-RFLP. Frequent shifts in composition were apparent across samples collected at approximately bi-weekly to monthly intervals. There were significant differences (ANOSIM, p < 0.05) in community composition between the Aransas and Mission River Estuary and Aransas Bay sites. Although the overall ANOSIM tests show significance between eukaryotic plankton communities monthly and between the bay water quality periods, none of the pairwise comparisons were significantly different. However, the ANOSIM R-statistic for the monthly pairwise comparisons displays a general increasing trend over time from sampling, further highlighting the dynamic nature of the microbial eukaryotic assemblage within sites.text
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Bae, Hanhong. "RFLP analysis of genetic variation in the laminated-root-rot fungal pathogen of conifers, Phellinus weirii /." 1992. http://hdl.handle.net/1957/10116.
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Wu, Hui-Ju, and 吳慧如. "Availability of Using RAPD(Random Amplified Polymorphic (DNA) Segments as Hybridization Probes in Sorghum RFLP analysis." Thesis, 1993. http://ndltd.ncl.edu.tw/handle/96177597261715815246.
Full textAbstract:
碩士國立臺灣大學
農藝學系
81
The molecular markers had been developed as a powerful tool for the studies in plant genetics and breeding. There are two types of molecular markers : one is restriction fragment length polymorphism (RFLP) marker, and the other is random amplified polymorphic DNA (RAPD) marker. In order to obtain the RFLP markers, it should take a long time and spend much resources to construct and screen a genetic library. However, since the polymerase chain reaction (PCR) technique was developed, it has revolutionized the RAPD marker production. RAPD markers are polymorphic DNA segments separated by gel electrophoresis after PCR amplification using random primers. Genetic analysis with RAPD markers is fast and involves no radioactivity and hybridization. For our current sorghum RFLP map construction, we had screened the informative probes from a PstI and a HpaII genomic libraries. In this study, we exploited the availability of using polymorphic DNA segments appeared in RAPD analysis as probes in RFLPs. Two sorghum parents, V-3 and V-160, were used as sources of DNA. We had screened out 192 amplified segments in 221 arbitrary 10-mer primers. And 142 of them had been used in RFLP analysis. Most of them were with a fragment size larger than 1.0 kb and shown to be multiple copies. There were 54 segments shown to be suitable as RFLP probes in these 142 segments. To explore the features of RFLP in sorghum, the relationship between the probability that a given enzyme detected polymorphism with a given probe in this study and the number of other enzymes detecting polymorphism with the same probe was plotted for regression. The result of regression suggested that insertion/ deletion was the major mechanism for generating RFLPs in the sorghum genome, however, the base replacement was not excluded.
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Chen, Hsiao-Jan, and 陳小然. "Identification of bovis group streptococci by PCR-RFLP based on groESL sequence and analysis of erythromycin-resistance determinant." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/59242363828570391423.
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碩士國立臺灣大學
醫事技術學研究所
93
Streptococcus bovis group is normal flora of the ruminant and human gut, but will also cause serious infections. Many reports have suggested a potential relationship between underlying infection with this organism and colon cancers. According to biochemical characteristics, S. bovis strains were divided into three biotypes, biotype I, Ⅱ/1 and Ⅱ/2. The clinical identification of bovis group depends on conventional methods or commercial rapid identification systems. But these two methods are time-consuming and not satisfactory. The diversity of biochemical characteristics of bovis group may lead to misidentification. Recently scientists have developed molecular identification based on 16S rRNA gene sequence. Limited to the highly conservation of 16S rRNA gene sequence, it is difficult to differentiate closely related species. Since the taxonomy of the species within bovis group has been proposed, the aim of this part of study was to develop a more rapid and reliable method for identification. In this study, we analyzed groESL sequences, which are ubiquitous and evolutionarily highly conserved, to understand the correlation between the phylogenetic evolution and the biotypes by PCR-direct sequencing method. The results revealed that isolates with the same biotype usually belong to the same genocluster. Then we designed specific PCR for the bovis group, and developed PCR-RFLP method to subsequently differentiate three biotypes. Resistance to erythromycin in S. bovis is mostly due to the target modification, which is mediated by erythromycin ribosome methylation (erm) that methylates 23S rRNA and induces ribosome modification. Because the target site of erythromycin is overlap with Lincosamide and Streptogramin B，ribosome modification can cause three antibiotics resistance known as macrolide-lincosamide-streptogramin B (MLSB) resistance. Expression of MLSB resistance can be either constitutive (cMLSB) or inducible (iMLSB). In previous studies, we found high rate of iMLSB strains in S. bovis. Detection of erythromycin resistance genes by PCR and sequence indicated that some iMLSB strains had erm(T) that is not reported in streptococci before. In this study, we analyzed the structures of erm(T) gene in these iMLSB strains. The SmaI PFGE fragments revealed the heterogeneity and not the expansion of a single clone. Furthermore, we have confirmed that the erm(T) resistance gene is located on chromosome.
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Yang, Chen-Chieh, and 楊鎮琾. "Studies of PCR-RFLP Analysis of Mitochondrial Gene on Identification of Species and Related Products of Cephalopod Species." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/96280413053034732919.
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碩士國立臺灣海洋大學
食品科學系
95
Cephalopoda is an important commercial catch in Taiwan. They play important roles not only in ecological area but in business sector. The distinctive morphological or physiological parameters provide the useful information to the blood relationship. The processing usually involved the separation of head, arms, and fins, evisceration, skinning, and freezing, which resulted in the change of morphological parameters. This fact makes it desirable to have methods for identifying cephalopod species, even in those cases when morphological characteristics have been removed. In order to establish the gene identification of cephalopod species, direct sequencing and the polymerase chain reaction – restriction fragment length polymorphism (PCR-RFLP) technology were used to determine the genetic variation of seven cephalopod species in this study. The gene probe was also developed to identify the cephalopod species of frozen, fresh, cooked and related products. The primers 16S L22/R21 were used to amplify partial mitochondrial gene of seven cephalopod species, including Suborder Oegopsida: Ommastrephes bartrami, Dosidicus gigas, Todarodes pacificus; Suborder Myopsida: Loligo chinensis, Sepioteuthis lessoniana; Order Sepiida: Sepia esculenta, and Order Octopoda: Octopus ocellatus. After analyzing the different sequences amplified with primers 16S L22/R21, two restriction enzymes including Ssp I and Ase I with specific cutting sites were used. The restriction enzyme Ssp I could differentiate the species of Ommastrephes bartrami, Dosidicus gigas, Sepioteuthis lessoniana, Sepia esculenta and Octopus ocellatus. Ase I could differentiate the species of Todarodes pacificus and Loligo chinensis. The polymorphic pattern in the DNA electrophoretic gel could support to identify these species precisely and quickly. Furthermore, applying above methods to identify the cephalopod species of frozen, cooked, sterilized and related products. The seven cephalopod meats were frozen at -20℃ for 3 months and heated at 100℃ for 30 minutes to obtain 14 samples. All samples were found to be amplified successfully. However when the cephalopod meats were sterilized at 121℃ for 15 minutes, DNA was degraded seriously and the samples could not be amplified. Besides, we analyzed 10 commercial cephalopod processed products sold in the market, of which 9 samples were determined successfully. The processed product of Sepia esculenta was failure in amplifying the target sequence. Therefore, this study provids useful technique to identify the species of cephalopod processed products.
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Chang, Chung-Yuh, and 張君玉. "Analysis of Genetic Diversity of Two Intersterility Groups of~u2 Ganoderma australe~u1 by RFLP and DNA sequencing." Thesis, 1996. http://ndltd.ncl.edu.tw/handle/60735454676811352442.
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碩士國立臺灣師範大學
生物學系
84
australe (Fr. ) Pat. was investigated by polymerase chain reaction,restriction fragment length polymorphism, and DNA sequencing.Two cultures of G. applanatum (Pers. ex Grau) Pat. CBS 187.31 and G. neo-japonicum Imaz. CCRC 36049 were selected for comparative studies. By using conserved primers complementary to rRNA genes of Saccharomyces cerevisiae, a 2.1 Kb fragment containing 17S rRNA gene and a 1.85 Kb fragment containing 5` half of 25S rRNA gene were efficiently amplified. Electrophoresis of HhaI, AvaII or HinfI digested PCR products produced restriction phenotypes. The two intersterility groups of G. arstrale can be differentiated by AvaII and HinfI restriction phenotypes. Detail mapping of restriction sites within rDNA locus by partial digestion and Southern hybridization reveals conserved and variable restriction sites for each species or group. The regions containing internal transcribed spacers were further amplified and subcloned into pGEM-T for DNA sequencing. Phylogenetic structures were constructed from analysis of ITS regions by PAUP and MEGA programs. Our data suggest the two groups of G. australe are genetically differentiated.
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Lin, Shin-Yuan, and 林欣緣. "Application of PCR-RFLP Analysis of Mitochondrial 16S Ribosomal RNA Gene on Identification of Species of Toxic Marine Snail." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/60937842898430264254.
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碩士國立臺灣海洋大學
食品科學系
94
Approximately 88,000 species of gastropod mollusk distribute in the world. However, several species of Naticidae, Nassariidae and Olividae have been detected to contain marine toxins, and outbreaks of food poisoning cases in Taiwan occurred. Species is usually identified by morphological characteristics of marine snail. But it is difficult to identify without shell. Recently, species identification is developed based on DNA analysis, especially the polymerase chain reaction-restriction fragment length polymorphism ( PCR-RFLP ). In order to identification of several toxic gastropod species in Taiwan, the direct sequencing and the PCR-RFLP technology were used to identify gastropod species in fresh, frozen, cooked ( 100℃, 30-60 min ) and steam sterilized ( 121℃, 15-30 min ) meat of Natica lineata, N. vitellus, Polinices didyma, P. tumidus,. Nassarius glans, N. papillosus, Niotha clathrata, Zeuxis scalaris, Oliva lignaria and O. reticulata. Judging from the data of this study, high molecular weight DNA（＞1,000 bp） was obtained from fresh, frozen and cooked meat, while relatively low molecular weight DNA was obtained from samples treated by high temperature. The PCR primers of 16S L22/R22 specifically amplify 383-396 bp of fragments from fresh, frozen and cooked meats; smaller fragments of 244-257 bp are used for steam sterilized meats amplified with primers of 16S LS22/RS22. After analyzing the different sequences amplified with primers 16S L22/R22 and 16S LS22/RS22, five restriction enzymes with specific cutting sites, including Apo I, Pac I, Hinc II, Acc I and Vsp I, were used. The restriction enzyme Apo I could differentiate the species of Natica lineata, N. vitellus, Polinices didyma and P. tumidus. Pac I, Hinc II and Acc I could differentiate the species of Nassarius glans, N. papillosus, Niotha clathrata and Zeuxis scalaris. Vsp I could differentiate the species of Oliva lignaria and O. reticulata. The polymorphic pattern in the DNA electrophoretic gel could support to identify these species precisely and quickly. By the same restriction enzyme could differentiate species of fresh, frozen, cooked and sterilized meats simultaneously. Furthermore, applying above method to identify the gastropod species in the residue of cooked gastropod meat from food poisoning incident in Hsiau Liouchiou Island, after shell identification, sequence comparing and PCR-RFLP analyzing, it indicated that the species of gastropod was Nassarius papillosus. Therefore, by this study could provide useful technique in identifying the gastropod species of different heating meat.
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Lin, Wen-Feng, and 林文風. "Application of PCR-RFLP Analysis of Mitochondrial Cytochrome b Gene on Identification of Species for Raw Material and Canned Products of Thunnus Tuna Species." Thesis, 2003. http://ndltd.ncl.edu.tw/handle/09959744551733678105.
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碩士國立海洋大學
食品科學系
91
Tuna is an important pelagic capture in Taiwan, the familiar species in Taiwan are bluefin tuna (Thunnus thynnus), albacore (T. alalunga), bigeye tuna (T. obesus) and yellowfin tuna (T. albacares). Tuna is a higher-price fish, and different species of tuna are quite different in cost. Bluefin tuna is the highest-price Thunnus species, and other raw fillets in the market are usually light meat, like bigeye tuna and yellowfin tuna. Traditionally, the raw material of canned tuna species is mainly white meat albacore. In order to establish the gene identification of fresh meat of familiar tuna species in Taiwan, in this study we used the directed sequencing and the polymerase chain reaction — restriction fragment length polymorphism (PCR-RFLP) technology to determine the genetic variation of four Thunnus species. Then we developed gene probes to identify the species of frozen, fresh and canned tuna meat. In the part of the diversity of sea areas and groups, T. thynnus was obtained from northern Pacific Ocean and northern Atlantic Ocean, and other three species were from northern Pacific Ocean, northern Atlantic Ocean and Indian Ocean. Judging from the data of this study, the 358 bp fragment of the mitochondrial cytochrome b gene in T. obesus was mostly different from other species, there was the highest divergence (4.235%). And the highest similarity was between T. thynnus and T. alalunga (98.687%∼99.674%). After analyzing the difference of sequence in four Thunnus species, we chose three restriction enzymes with specific cutting sites, including Bsp1286 I, Hinc II and Rsa I. The restriction enzyme Bsp1286 I cleaved the 358 bp fragment to 283 bp and 75 bp in T. thynnus, and there was no any cutting site in other three species. Hinc II could cleave the 358 bp fragments of T. thynnus, T. alalunga and T. albacares to separate into 198 bp, 150 bp and 10 bp, but there was no cutting site in T. obesus. And then, Rsa I cleaved tuna fishes both T. thynnus and T. alalunga into fragments of 256 bp and 102 bp, it was different from the fragments of 284 bp and 74 bp in T. albacares and T. obesus. The polymorphic pattern in the DNA electrophoretic gel could identify four fresh Thunnus species precisely and quickly. Furthermore, we analyzed the 12 raw fillets of tuna sold in the market and found that there was no case to personate high-price T. thynnus by other tuna species. In the part of canned product, we designed 7 pairs of primer for PCR amplifying. We added different sauces into tuna meat and then heated at 121℃ for 15 min and 115℃ for 30 min to simulate the process of canning. However, the DNA was degraded seriously, there were only two pairs of primer CbBRs126L/H and CbHi146L/H could successfully amplify the PCR products, short fragments of 126 bp and 146 bp, respectively. Then we used three restriction enzymes Bsp1286 I, Hinc II and Rsa I to analyze the cutting sites of procssed products and could differentiate the diversity between four species by the DNA electrophoretic map. Last, we analyzed 10 commercial canned products sold in the market, except 2 samples were failure in amplifying the target sequence, other 8 samples including 6 cases of T. albacares and 2 cases of T. alalunga were determined successfully. Therefore, this study could provide useful and academic technique, in identifying the species of processed tuna meats.
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Kernaghan, Shaun. "Characterization of the Bacterial Communities of the Tonsil of the Soft Palate of Swine." Thesis, 2013. http://hdl.handle.net/10214/7750.
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Terminal restriction fragment length polymorphism (T-RFLP) analysis and pyrosequencing were used to characterize the microbiota of the tonsil of the soft palate of 126 unfit and 18 healthy pigs. The T-RFLP analysis method was first optimized for the study of the pig tonsil microbiota and the data compared with culture-based identification of common pig pathogens. Putative identifications of the members of the microbiota revealed that the phyla Firmicutes, Proteobacteria and Bacteroidetes were the most prevalent. A comparison of the T-RFLP analysis results grouped into clusters to clinical conditions revealed paleness, abscess, PRRS virus, and Mycoplasma hyopneumoniae to be significantly associated with cluster membership. T-RFLP analysis was also used to select representative tonsil samples for pyrosequencing. These studies confirmed Actinobacteria, Bacteroidetes, Firmicutes, Fusobacteria, and Proteobacteria to be the core phyla of the microbiota of the tonsil of the soft palate of pigs.OMAFRA Animal Health Strategic Investment
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Johnson, Meredith Christina. "Understanding rumen fermentation i. effect of high DHA algal oil on microbial biohydrogenation and ii. monitoring microbial shifts in response to antibiotics and oil using T-RFLP analysis /." 2007. http://www.lib.ncsu.edu/theses/available/etd-06272007-103423/unrestricted/etd.pdf.
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SEPEHRI, SHADI. "Microbial etiology of Inflammatory Bowel Disease: Microbial diversity and the role of Escherichia coli." 2010. http://hdl.handle.net/1993/3977.
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Inflammatory bowel disease (IBD), comprises Crohn’s disease (CD) and ulcerative colitis (UC), and is a chronic relapsing inflammation of gastrointestinal tract without any known cause or cure. Currently, it is accepted that IBD is a result of a dysfunctional immune response to commensal bacteria in a genetically susceptible host, and that environmental factors can trigger the onset or reactivation of the disease. This thesis considers the possibility of a specific pathogenic agent as well as an imbalance in the composition of the normal microflora in the pathogenesis of IBD. Gut biopsy tissues were taken from a population-based case-control tissue bank held at the University of Manitoba. Automated ribosomal intergenic spacer analysis (ARISA) and terminal restriction fragment length polymorphisms (T-RFLP) were employed to assess the diversity of gut microbiota. The phylogenetic, virulence and biochemical characteristics of Escherichia coli isolated from IBD biopsies were examined using multi-locus sequence typing (MLST), DNA microarray technology and API 20E system. Utilizing ARISA and T-RFLP, a remarkable increase in the order of unclassified Clostridia was detected in inflamed tissues, particularly in CD patients (P < 0.05). Moreover, species richness and diversity were the highest in non-inflamed IBD biopsies. Culture-based quantification detected a significantly higher number of E. coli in IBD tissues (P < 0.05). Phylogenetic analysis revealed the tendency of E. coli isolated from IBD patients to be grouped into separate clonal clusters based on their allelic profiles (P = 0.02). A link was detected between uropathogenic E. coli (UPEC) CFT073 and strains isolated from IBD, with regards to gene distribution and virulence, using microarray technology. Amino acid substitutions N91S and S99N in FimH, the adhesive subunit of E. coli type I fimbria, were significantly associated to IBD (P < 0.05). This study demonstrated an increase in the microbial diversity of non-inflamed IBD tissues and suggested a recruitment phase of bacterial adherence and colonization, before the inflammation sets in. Furthermore, E. coli isolated from IBD tissues were distinct from commensal strains in both clonal and virulence characteristics and shared remarkable traits with extraintestinal pathogenic E. coli. Features involved in bacterial adhesion to epithelial cells may hold the key to E. coli pathogenesis in IBD.
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Bark, Hur Ockyung. "RFLP analyses of the chloroplast and nuclear genomes to establish cytoplasms, demonstrate interspecific DNA transfer, and estimate relationships among bulb-onion populations." 1993. http://catalog.hathitrust.org/api/volumes/oclc/30799736.html.
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