Relevant bibliographies by topics / Multiplex amplification

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  1. Journal articles
  2. Dissertations / Theses
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  5. Conference papers
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Academic literature on the topic 'Multiplex amplification'

Author: Grafiati
Published: 4 June 2021
Last updated: 19 February 2022

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Journal articles on the topic "Multiplex amplification"

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Jeuken, Judith, Sandra Cornelissen, Sandra Boots-Sprenger, Sabine Gijsen, and Pieter Wesseling. "Multiplex Ligation-Dependent Probe Amplification." Journal of Molecular Diagnostics 8, no. 4 (September 2006): 433–43. http://dx.doi.org/10.2353/jmoldx.2006.060012.

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Broude, Natalia E., Kristina Driscoll, and Charles R. Cantor. "High-Level Multiplex DNA Amplification." Antisense and Nucleic Acid Drug Development 11, no. 5 (October 2001): 327–32. http://dx.doi.org/10.1089/108729001753231704.

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Römpler, Holger, Paul H. Dear, Johannes Krause, Matthias Meyer, Nadin Rohland, Torsten Schöneberg, Helen Spriggs, Mathias Stiller, and Michael Hofreiter. "Multiplex amplification of ancient DNA." Nature Protocols 1, no. 2 (July 13, 2006): 720–28. http://dx.doi.org/10.1038/nprot.2006.84.

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Cave, Christopher A., Kristen Hancock, and James W. Schumm. "Principles of STR multiplex amplification." Forensic Science International: Genetics Supplement Series 1, no. 1 (August 2008): 102–4. http://dx.doi.org/10.1016/j.fsigss.2007.10.016.

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Millholland, J., S. G. Patel, C. A. Fernandez, and A. P. Shuber. "Development of a real-time multiplex PCR assay for the detection of FGFR3 mutations in urine." Journal of Clinical Oncology 29, no. 7_suppl (March 1, 2011): 271. http://dx.doi.org/10.1200/jco.2011.29.7_suppl.271.

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271 Background: We have recently reported the development of a Multi-Analyte Diagnostic Readout (MADR) non-invasive assay using urinary matrix metalloproteinases (MMPs) and FGFR3 as triage monitors in high-risk bladder cancer populations. This concept combines the marker performance characteristics of protein and DNA biomarkers into one assay for optimal performance. Eight common FGFR3 mutations in 3 exons have been associated with bladder cancer. Analysis of mutational status for each single mutation required 8 amplification steps, which were costly and time consuming. We have now developed a real-time multiplexed FGFR3 assay, generating a cost-effective, clinically applicable assay for the detection of FGFR3 mutations in urine. Methods: Our approach involves a two-step PCR amplification process. The initial round generates exon specific PCR products, which are then used as template for real-time PCR mutation detection utilizing locked nucleic acid (LNA) oligonucleotides. The LNA suppress wild-type DNA amplification. To convert our existing FGFR3 assay to a multiplex format, primary amplifications of exons 7, 10, and 15 were combined into a single real-time PCR assay for exon specific amplification and DNA quantitation. The LNA-mediated mutation detection was then converted from 4 reactions to 2 duplex amplifications. All multiplex assays were carried out on the Roche LC 480 real-time PCR platform. Results: To validate the new multiplex format, FGFR3 multiplex analysis was performed on DNA isolated from 50 Ta stage bladder tumors. FGFR3 mutations were detected in 90% (48/50) of the tumors. To directly compare performance with single mutation analysis, 40 urine samples previously analyzed using the singleplex format were again tested using the multiplex FGFR3 assay. 100% concordance was seen between the two assay formats. Conclusions: By multiplexing the FGFR3 mutation analysis we reduced the number of amplification steps, improving assay turnaround time and throughput, without compromising assay performance. The FGFR3 multiplex analysis provides a robust, cost-effective DNA assay that in combination with MMP protein analysis delivers a clinically applicable assay with optimal performance. [Table: see text]
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Moelans, Cathy B., Hanneke N. Monsuur, Johannes H. de Pinth, Remco D. Radersma, Roel A. de Weger, and Paul J. van Diest. "ESR1 Amplification is Rare in Breast Cancer and is Associated with High Grade and High Proliferation: A Multiplex Ligation-Dependent Probe Amplification Study." Analytical Cellular Pathology 33, no. 1 (2010): 13–18. http://dx.doi.org/10.1155/2010/619180.

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Background: Expression of estrogen receptor alpha (ERα) is predictive for endocrine therapy response and an important prognostic factor in breast cancer. Overexpression of ERα can be caused by estrogen receptor 1 (ESR1) gene amplification and was originally reported to be a frequent event associated with a significantly longer survival for ER-positive women treated with adjuvant tamoxifen monotherapy, which was however questioned by subsequent studies.Methods: This study aimed to reanalyze the frequency of ESR1 amplification by multiplex ligation-dependent probe amplification (MLPA) and fluorescence in situ hybridisation (FISH), and to assess clinicopathologic correlations. MLPA was performed in a group of 135 breast cancer patients, and gains/amplifications were subjected to FISH.Results: True ESR1 amplification by MLPA was rare (2%) and only 6% more patients showed a modest gain of ESR1. All MLPA-detected ESR1 amplifications and nearly all ESR1 gains were also FISH amplified and gained, but not all FISH amplifications/gains were MLPA amplified/gained, leading to an overall concordance of only 60% between both techniques. All 3 MLPA and FISH ESR1 amplified cases had high ERα expression, but there was no obvious correlation between ESR1 gain and ER status by IHC. ESR1 gains/amplifications were not associated with HER2 gain/amplification, but seemed to be associated with older age. Surprisingly, ESR1 gain/amplification was not associated with low grade as reported previously, but correlated with high grade and high proliferation. Furthermore, ESR1 gain/amplification by MLPA was not associated with nodal status or tumor size (pT status).Conclusion: ESR1 amplification as detected by MLPA is rare in breast cancer, and seems to be associated with high ERα expression, high age, high grade and high proliferation. This study confirms previous studies that showed differences in the ESR1 amplification frequencies detected by different techniques.
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Minarikova, Petra, Lucie Benesova, Tereza Halkova, Barbora Belsanova, Inna Tuckova, Frantisek Belina, Ladislav Dusek, Miroslav Zavoral, and Marek Minarik. "Prognostic Importance of Cell Cycle Regulators Cyclin D1 (CCND1) and Cyclin-Dependent Kinase Inhibitor 1B (CDKN1B/p27) in Sporadic Gastric Cancers." Gastroenterology Research and Practice 2016 (2016): 1–8. http://dx.doi.org/10.1155/2016/9408190.

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Background. Gastric cancer is known for a notable variety in the course of the disease. Clinical factors, such as tumor stage, grade, and localization, are key in patient survival. It is expected that molecular factors such as somatic mutations and gene amplifications are also underlying tumor biological behavior and may serve as factors for prognosis estimation.Aim. The purpose of this study was to examine gene amplifications from a panel of genes to uncover potential prognostic marker candidates.Methods. A panel of gene amplifications including 71 genes was tested by multiplex ligation-dependent probe amplification (MLPA) technique in 76 gastric cancer samples from a Caucasian population. The correlation of gene amplification status with patient survival was determined by the Kaplan-Meier method.Results. The amplification of two cell cycle regulators,CCND1andCDKN1B, was identified to have a negative prognostic role. The medial survival of patients with gastric cancer displaying amplification compared to patients without amplification was 192 versus 725 days forCCND1(P=0.0012) and 165 versus 611 days forCDKN1B(P=0.0098).Conclusion. Gene amplifications ofCCND1andCDKN1Bare potential candidates to serve as prognostic markers for the stratification of patients based on the estimate of survival in the management of gastric cancer patients.
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Søe, Martin Jensen, and Peter Warthoe. "RT-isoPCR: nested, high multiplex mRNA amplification." Analyst 138, no. 20 (2013): 5871. http://dx.doi.org/10.1039/c3an00803g.

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Meuzelaar, Linda Strömqvist, Owen Lancaster, J. Paul Pasche, Guido Kopal, and Anthony J. Brookes. "MegaPlex PCR: a strategy for multiplex amplification." Nature Methods 4, no. 10 (September 16, 2007): 835–37. http://dx.doi.org/10.1038/nmeth1091.

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Talavera Vargas-Machuca, Sergio, Ismenia Gamboa Oré, Francia Huamán Dianderas, Ricardo Fujita Alarcón, María Luisa Fajardo Loo, and María Luisa Guevara Gil. "Diagnóstico molecular de síndrome de Smith-Magenis por MLPA (Multiplex Ligation-dependent Probe Amplification)." Horizonte Médico (Lima) 17, no. 3 (June 30, 2017): 73–78. http://dx.doi.org/10.24265/horizmed.2017.v17n3.12.

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Dissertations / Theses on the topic "Multiplex amplification"

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Zabre, Sidkièta. "Amplification non-linéaire d’un multiplex de porteuses modulées à fort facteur de crête." Rennes 1, 2007. http://www.theses.fr/2007REN1S014.

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Au nombre des difficultés de mise en œuvre de la Radio Logicielle (RL) réside le problème de l’amplification non linéaire des signaux à fort variation de puissance. Il est connu, que les amplificateurs de puissance présentent des caractéristiques non linéaires; et si tôt qu'un signal radio fréquence à fortes fluctuations d'enveloppe doit être amplifié, des distorsions néfastes viennent perturber de façon non négligeable les données utiles entraînant une dégradation des performances de communications. Pour apporter une solution au problème posé, nous avons scindé celui-ci en deux parties. La première partie à consister à modéliser les signaux RL puis à caractériser la notion de variation de puissance dans le contexte RL. Nous avons défini un paramètre, le Power Ratio (PR) qui traduit cette notion de variation de puissance. Ensuite nous avons analysé le PR du cas le plus simple (monoporteuse) au cas le plus complexe (multi-standards). Cette analyse nous a conduit à proposer une nouvelle vision du Power Ratio dans le cas multiporteuse OFDM. Une analyse fortement inspirée du cas multiporteuse est proposée dans le contexte Radio Logicielle. Dans la seconde partie de notre étude nous avons proposé le concept novateur de «méthode des porteuses fantômes C-SOCP » pour réduire le PR. Cette méthode s’applique à tout type de signal. De plus elle est à compatibilité descendante et ne dégrade point le taux d’erreur binaire. Nous avons alors modélisé le problème de réduction du PR sous forme d’un problème d’optimisation convexe plus précisément sous forme d’un Second Order Cone Programming (SOCP). Nous ajoutons à notre modélisation initiale, des contraintes soit pour contrôler la puissance additionnelle transmise, soit pour respecter le gabarit du spectre spécifié dans les standards.
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Suemasu, Cintia Natsumi. "Caracterização dos genótipos da talassemia alfa delecional por MLPA (Multiplex Ligation-Dependent Probe Amplification)." [s.n.], 2010. http://repositorio.unicamp.br/jspui/handle/REPOSIP/308947.

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Orientador: Maria de Fátima Sonati
Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas
Made available in DSpace on 2018-08-16T20:41:19Z (GMT). No. of bitstreams: 1 Suemasu_CintiaNatsumi_M.pdf: 6481597 bytes, checksum: f9093ca98b4efb60bfe4f81a7e80efee (MD5) Previous issue date: 2010
Resumo: A talassemia alfa (a) constitui um grupo de doenças hereditárias causadas pela redução ou ausência da síntese das cadeias 'alfa' da hemoglobina. Os genes responsáveis pela codificação dessas cadeias são duplicados ('alfa' 2 e 'alfa'1) e estão localizados em um agrupamento gênico ou cluster, situado próximo ao telômero do cromossomo 16 (16p13.3). As deleções são as principais causas da doença, podendo afetar um ou ambos os genes 'alfa' do cromossomo (formas 'alfa' + e 'alfa' 0, respectivamente). Nos últimos anos, várias técnicas foram desenvolvidas para identificar as deleções envolvendo o cluster dos genes a no cromossomo 16. A gap- PCR, o Southern blot e o FISH são comumente aplicados com esta finalidade; entretanto, muitas deleções ainda não são detectadas utilizando-se essas estratégias metodológicas. Em 2005, uma técnica simples e adequada a análises quantitativas rápidas, o Multiplex Ligation-dependent Probe Amplification (MLPA), foi adaptada ao diagnóstico das hemoglobinopatias. Esta técnica é baseada na ligação e na amplificação, em reação de PCR, de dois oligonucleotídeos (sondas) que se hibridizam de forma adjacente ao DNA alvo. Cada oligonucleotídeo é projetado para gerar um produto de comprimento único e, por possuírem terminações comuns, todas as sondas podem ser amplificadas utilizando um único par de iniciadores. Por fim, o uso de fluorocromos permite a separação dos fragmentos, que foram gerados pela amplificação das sondas, em sistema de seqüenciamento por capilaridade. No presente estudo nós utilizamos o MLPA para investigar as bases moleculares da talassemia alfa em nove pacientes não relacionados, cinco deles com Doença da Hb H, cujos diagnósticos moleculares não puderam ser esclarecidos pelas técnicas usuais, incluindo o seqüenciamento de DNA. No total, foram utilizadas 44 sondas, distribuídas desde o telômero ao gene MSLN, cobrindo uma região de cerca de 800 kb, na região 16p13.3. Oito diferentes deleções foram detectadas. Enquanto quatro delas comprometem uma região de grande extensão que inclui todo o locus a e seu elemento regulatório, com tamanhos mínimos de 240 kb, 470 kb, 500 kb e 720 kb, respectivamente (posições 97000-334571, 97000-570532, 97000-602492 and 97000- 816477 do UCSC Genome Browser, Fevereiro, 2009, respectivamente), quatro outras encontram-se limitadas às regiões que contém o elemento regulatório, deixando os genes a intactos, porém sem expressão. Essas últimas apresentam tamanhos mínimos de 0.4 kb, em um dos casos, e de 95 kb a 100 kb nos demais (posições entre 163464-163904, 97000- 193847, 97000-202417 e 97000-202417 do UCSC Genome Browser, Fevereiro, 2009, respectivamente). Em um dos pacientes com fenótipo talassêmico heterozigótico foi demonstrada a presença de uma quadruplicação de cerca de 230 kb de DNA, incluindo todo o cluster a e seu elemento regulatório (97000 a 231370 do UCSC Genome Browser, Fevereiro, 2009). Este estudo representa o primeiro no Brasil a empregar o método de MLPA na caracterização das bases moleculares da talassemia a. A ampla diversidade de alterações identificadas enfatiza a necessidade de se investigar todos os casos cujos valores hematológicos revelem hipocromia e microcitose, na ausência de deficiência de ferro e níveis normais de HbA2
Abstract: Alpha (a) thalassemia is an inherited hemoglobin disorder characterized by reduction or absence of the a-globin chain synthesis. These chains are encoded by duplicated genes ('alpha' 2 and 'alpha' 1) that lie in a gene cluster near the telomeric end of the short arm of chromosome 16 (16p13.3). Deletions are the major molecular cause of the disease and may affect one or both 'alfa genes in the chromosome (the 'alpha' + and 'alpha' 0 forms, respectively). In recent years several techniques have been developed to identify deletions involving the 'alpha' -globin cluster in chromosome 16. The molecular tests commonly used to identify these deletions are gap- PCR, Southern Blot analysis and FISH. However, several thalassemia deletions remain uncharacterized by these methods. In 2005 a simple technique suitable for rapid quantitative analysis ¾ multiplex ligationdependent probe amplification (MLPA) ¾ was adapted for the diagnosis of thalassemias. The approach is based on the ligation and PCR amplification of two adjacently hybridizing oligonucleotides (probes). Each oligonucleotide pair is designed to give a product of unique length, and by using common ends all the probes can be amplified with one primer pair. Finally, the use of a fluorescent label allows the probe to be separated in a capillary sequencing system. In this study we used MLPA to investigate the molecular basis of thalassemia in nine unrelated patients (five with Hb H disease) in whom the molecular causes of the disease could not be detected by sequence analysis or other conventional techniques. In total, 44 probe pairs were used, covering approximately 800 kb from the telomere to the MSLN gene in the 16p13.3 region. Eight different deletions were detected. While four of them affect a large region including the entire ? -globin gene locus and its upstream regulatory element, spanning genomic regions of at least 240 kb, 470 kb, 500 kb and 720 kb (positions 97000-334571, 97000-570532, 97000-602492 and 97000-816477 of the UCSC Genome Browser, February, 2009, respectively). The other four deletions were limited to a region that contains the upstream regulatory element; the ? -globin genes, although intact, are therefore not expressed. These deletions span a region of at least 0.4 kb in one case and from 95 kb to 100 kb in the other cases (positions 163464-163904, 97000- 193847, 97000-202417 and 97000-202417 of the UCSC Genome Browser, February, 2009, respectively). One carrier who was suspected of having heterozygous ? -thalassemia showed a segmental quadruplication spanning about 230 kb and including the a cluster and upstream regulatory element (97000-231370 do UCSC Genome Browser, February, 2009). This study is the first in Brazil to use the MLPA method to characterize thalassemias. The variety of rearrangements identified highlights the need to investigate all cases presenting with microcytosis and hypochromia without iron deficiency or elevated Hb A2 levels
Mestrado
Ciencias Biomedicas
Mestre em Ciências Médicas
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Ereku, Luck Tosan. "Design of microfluidic multiplex cartridge for point of care diagnostics." Thesis, Brunel University, 2017. http://bura.brunel.ac.uk/handle/2438/15331.

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A simple, but innovative microfluidic Lab-on-a-chip (LOC) device which is broadly applicable in point of care diagnostics of biological pathogens was designed, fabricated and assembled utilising explicit microfluidic techniques. The purpose of this design was to develop a cartridge with the capability to perform multiplex DNA amplification reactions on a single device. To achieve this outcome, conventional laboratory protocols for sample preparation; involving DNA extraction, purification and elution were miniaturized to suit this lab-on-a-chip device of 75mm X 50mm cross-sectional area. The extraction process was carried out in a uniquely designed microchamber embedded with chitosan membrane that binds DNA at pH 5.0 and elutes when a different solution at pH 9.0 flows through. Likewise, purification protocol that occurs in the designed waste reservoir is very significant in biomedical field because it is concerned with waste treatment and cartridge disposability, was performed with a super absorbent powder that converts liquid to a gel like substance. This powder is known as sodium polyacrylate, which is also they treated with anti-bacterial chemicals to prevent environmental contamination. Furthermore, this process also employed the use of a passive valve for a precise fluid handling operation involving flow regulation from extraction to waste reservoir. In order to achieve the intended multiplexing function a multiplexer was created to distribute flow simultaneously through a bifurcated network of channels connected to six similar amplification microchambers. Prior to fabrication, computational fluid dynamics (CFD) simulation was utilized at flowrates less than 10μL/s as the means to test the effectiveness of each design components and also to specifically deduct empirical values that can be analyzed to improve or understand the relationship between the fluid and geometrical constraints of the microfluidic modular elements. The device produced was a hybrid cartridge composed of PDMS and glass which is the most widely used materials microfluidics research due to their low cost and simplicity of fabrication by soft lithography technique. The choice of material also took into account the various physical and chemical properties advantages and disadvantages in their bio-medical applications. Such properties include but not limited to surface energy that determines the wetting fluid characteristics, biocompatibility, optical transparency. Subsequently, after a prototype cartridge was developed fluid flow experimentation using liquid coloured dye was used on the fully fabricated cartridge to test the efficacy of its microfluidic functionalities before expensive DNA amplification reagents were utilised at similar flowrates to the CFD simulations. This gave rise to comparison between similar and dissimilar flow Peculiarities in the microfluidic circuit of both experiments. The final experiment was performed with the aid of a recent molecular technique in DNA amplification known as of RPA kit (recombinase polymerase amplification reaction). It involved performing two main reaction experiments; first, was the positive experiment that bears the sample DNA and the latter, negative that served as the control without DNA. In the end, quantitative analysis of results was done using an agarose gel that showed 143 base pairs, for the positive samples, thus validating the amplification experiment.
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蘇昭燕 and Chiu-yin So. "Molecular characterization of large deletions in beta globin gene cluster using multiplex ligation-dependent probe amplification." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2008. http://hub.hku.hk/bib/B40738164.

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So, Chiu-yin. "Molecular characterization of large deletions in beta globin gene cluster using multiplex ligation-dependent probe amplification." Click to view the E-thesis via HKUTO, 2008. http://sunzi.lib.hku.hk/hkuto/record/B40738164.

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Cohn, Dianne Marie. "Utilizing multiplex ligation-dependent probe amplification to detect novel x-linked microduplications which cause intellectual disability." Connect to this title online, 2008. http://etd.lib.clemson.edu/documents/1239894829/.

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Huamán, Dianderas Francia Del Pilar. "Diagnóstico molecular de pacientes con retraso mental idiopático usando la técnica Multiplex Ligation Probe Amplification (MLPA)." Bachelor's thesis, Universidad Nacional Mayor de San Marcos, 2016. https://hdl.handle.net/20.500.12672/5468.

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Detecta las mutaciones de tipo deleción y/o duplicación causantes de RM idiopático síndrómico o no síndrómico en diferentes genes y regiones, por medio de la técnica MLPA, siguiendo los pasos propuestos por GIRMOGEN 2012. El lugar de ejecución fue el Centro de Genética y Biología Molecular (CGBM) del Instituto de Investigación de la Facultad de Medicina Humana de la Universidad San Martín de Porres. El material biológico consistió en una muestra de 3 a 4 mL sangre periférica, en tubos vacutainer con EDTA, de 50 pacientes diagnosticados clínicamente con retraso mental provenientes del Servicio de Genética - Departamento de Especialidades Médicas del Hospital Edgardo Rebagliati Martins – EsSalud.
Tesis
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Claesson, Cim. "Is Multiplex Ligation-dependent Probe Amplification a good method for screening formalin fixed paraffin embedded neuroblastoma tumors?" Thesis, Högskolan i Skövde, Institutionen för vård och natur, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:his:diva-5442.

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Neuroblastoma is one of the most enigmatic solid tumors for scientists and pediatric oncologists. Neuroblastoma is primary a childhood form of cancer, consisting of neuroectodermal cells that originate from the neural crest and is destined for the  adrenal medulla and sympathetic nervous system. The Neuroblastoma group at The University of Gothenburg received formalin-fixed paraffin-embedded  tumor samples from Vietnam and this project was to examine if the quality of the DNA from, is good enough to run comparative genome hybridization array experiment  on by using a cheaper technique Multiplex  Ligation-dependent Probe Amplification   technique. Multiplex Ligation-dependent Probe Amplification (MRC Holland) is a multiplex PCR method that can detect abnormalities such as deletions and amplifications. By using probes consisting of one short synthetic arm with a PCR primer sequence Y at the 3´end, and one long probe with a stuffer sequence, and a PCR primer sequence X at the 5´end that can hybridize and ligate. If these probes ligate it is possible to amplify them by PCR just using specific primers for the X and Y sequences. The resulting amplification products can then be analyzed bycapillary electrophoresis. These patient that the DNA was derived from had all stage 4  neuroblastoma, and that is why they all present many aberrations. Among the fascinating data from this experiment, there are many patients with both 11q  deletions and has an extreme amplification of MYCN. In Sweden only a few cases has been discovered. In this material even though all patients are stage 4 patients, 16 have this combination.
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Sandberg, Julia. "Massively parallel analysis of cells and nucleic acids." Doctoral thesis, KTH, Genteknologi, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-45671.

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Recent proceedings in biotechnology have enabled completely new avenues in life science research to be explored. By allowing increased parallelization an ever-increasing complexity of cell samples or experiments can be investigated in shorter time and at a lower cost. This facilitates for example large-scale efforts to study cell heterogeneity at the single cell level, by analyzing cells in parallel that also can include global genomic analyses. The work presented in this thesis focuses on massively parallel analysis of cells or nucleic acid samples, demonstrating technology developments in the field as well as use of the technology in life sciences. In stem cell research issues such as cell morphology, cell differentiation and effects of reprogramming factors are frequently studied, and to obtain information on cell heterogeneity these experiments are preferably carried out on single cells. In paper I we used a high-density microwell device in silicon and glass for culturing and screening of stem cells. Maintained pluripotency in stem cells from human and mouse was demonstrated in a screening assay by antibody staining and the chip was furthermore used for studying neural differentiation. The chip format allows for low sample volumes and rapid high-throughput analysis of single cells, and is compatible with Fluorescence Activated Cell Sorting (FACS) for precise cell selection. Massively parallel DNA sequencing is revolutionizing genomics research throughout the life sciences by constantly producing increasing amounts of data from one sequencing run. However, the reagent costs and labor requirements in current massively parallel sequencing protocols are still substantial. In paper II-IV we have focused on flow-sorting techniques for improved sample preparation in bead-based massive sequencing platforms, with the aim of increasing the amount of quality data output, as demonstrated on the Roche/454 platform. In paper II we demonstrate a rapid alternative to the existing shotgun sample titration protocol and also use flow-sorting to enrich for beads that carry amplified template DNA after emulsion PCR, thus obtaining pure samples and with no downstream sacrifice of DNA sequencing quality. This should be seen in comparison to the standard 454-enrichment protocol, which gives rise to varying degrees of sample purity, thus affecting the sequence data output of the sequencing run. Massively parallel sequencing is also useful for deep sequencing of specific PCR-amplified targets in parallel. However, unspecific product formation is a common problem in amplicon sequencing and since these shorter products may be difficult to fully remove by standard procedures such as gel purification, and their presence inevitably reduces the number of target sequence reads that can be obtained in each sequencing run. In paper III a gene-specific fluorescent probe was used for target-specific FACS enrichment to specifically enrich for beads with an amplified target gene on the surface. Through this procedure a nearly three-fold increase in fraction of informative sequences was obtained and with no sequence bias introduced. Barcode labeling of different DNA libraries prior to pooling and emulsion PCR is standard procedure to maximize the number of experiments that can be run in one sequencing lane, while also decreasing the impact of technical noise. However, variation between libraries in quality and GC content affects amplification efficiency, which may result in biased fractions of the different libraries in the sequencing data. In paper IV barcode specific labeling and flow-sorting for normalization of beads with different barcodes on the surface was used in order to weigh the proportion of data obtained from different samples, while also removing mixed beads, and beads with no or poorly amplified product on the surface, hence also resulting in an increased sequence quality. In paper V, cell heterogeneity within a human being is being investigated by low-coverage whole genome sequencing of single cell material. By focusing on the most variable portion of the human genome, polyguanine nucleotide repeat regions, variability between different cells is investigated and highly variable polyguanine repeat loci are identified. By selectively amplifying and sequencing polyguanine nucleotide repeats from single cells for which the phylogenetic relationship is known, we demonstrate that massively parallel sequencing can be used to study cell-cell variation in length of these repeats, based on which a phylogenetic tree can be drawn.
QC 20111031
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Werne, Solnestam Beata. "Interpreting the human transcriptome." Doctoral thesis, KTH, Genteknologi, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-158320.

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The human body is made of billions of cells and nearly all have the same genome. However, there is a high diversity of cells, resulted from what part of the genome the cells use, i.e. which RNA molecules are expressed. Rapid advances within the field of sequencing allow us to determine the RNA molecules expressed in a specific cell at a certain time. The use of the new technologies has expanded our view of the human transcriptome and increased our understanding of when, where, and how each RNA molecule is expressed. The work presented in this thesis focuses on analysis of the human transcriptome. In Paper I, we describe an automated approach for sample preparation. This protocol was compared with the standard manual protocol, and we demonstrated that the automated version outperformed the manual process in terms of sample throughput while maintaining high reproducibility. Paper II addresses the impact of nuclear transcripts on gene expression. We compared total RNA from whole cells and from cytoplasm, showing that transcripts with long, structured 3’- and 5’-untranslated regions and transcripts with long protein coding sequences tended to be retained in the nucleus. This resulted in increased complexity of the total RNA fraction and fewer reads per unique transcript. Papers III and IV describe dynamics of the human muscle transcriptome. For Paper III, we systematically investigated the transcriptome and found remarkably high tissue homogeneity, however a large number of genes and isoforms were differentially expressed between genders. Paper IV describes transcriptome differences in response to repeated training. No transcriptome-based memory was observed, however a large number of isoforms and genes were affected by training. Paper V describes a transcript profiling protocol based on the method Reverse Transcriptase Multiplex Ligation-dependent Probe Amplification. We designed the method for a few selected transcripts whose expression patterns are important for detecting breast cancer cells, and optimized the method for single cell analysis. We successfully detected cells in human blood samples and applied the method to single cells, confirming the heterogeneity of a cell population.
Människokroppen är uppbyggd av miljarder celler och nästan alla innehåller samma arvsmassa. Trots detta finns det många olika celler med olika funktioner vilket är en följd av vilken del av arvsmassan som cellerna använder, dvs vilka RNA-molekyler som finns i varje cell. Den snabba utvecklingen av sekvenseringstekniker har gjort det möjligt att studera när, var och hur varje RNA-molekyl är uttryckt och att få en djupare förståelse för hur människans celler fungerar. Arbetet som presenteras i denna avhandling fokuserar på analys av RNA-molekyler i människans celler. I artikel I beskriver vi en automatiserad metod för att förbereda cellprov för RNA-sekvensering. Det automatiserade protokollet jämfördes med det manuella protokollet, och vi visade att det automatiserade protokollet överträffade det manuella när det gällde provkapacitet samtidigt som en höga reproducerbarheten behölls. I artikel II undersökte vi effekterna som RNA-molekyler från en del av cellen (cellkärnan) har på den totala mängden uttryckta RNA-molekyler. Vi jämförde RNA från hela cellen och från en del av cellen (cytoplasman) och visade att RNA-molekyler med långa och strukturerade 3'- och 5'-otranslaterade regioner och RNA-molekyler med långa proteinkodande sekvenser tenderade att hållas kvar i cellkärnan till en högre grad. Detta resulterade i en ökad komplexitet av RNA-molekylerna i hela cellen, medan vi i cytoplasma-fraktionen lättare kunde hitta de korta och svagt uttryckta RNA-molekyler. I Artikel III och IV studerar vi RNA-molekyler i människans skelettmuskler. I artikel III visar vi att andelen RNA-molekyler uttryckta i skelettmuskler är väldigt lika mellan muskler och mellan olika personer, men att ett stort antal RNA-molekyler var uttryckta i olika nivåer hos kvinnor och män. Artikel IV beskriver RNA-nivåer som svar på upprepade perioder av uthållighetsträning. Artikel V beskriver en metod för att studera ett fåtal utvalda RNA-molekyler. Vi valde RNA-molekyler vars uttryck är viktigt vid analys av bröstcancerceller, och optimerade metoden för analys av enskilda celler. Vi analyserade cancerceller från blodprov och använde metoden för att titta på RNA-nivåer i enskilda celler från en grupp av celler och visade på skillnader i RNA-nivåer inom gruppen.

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Books on the topic "Multiplex amplification"

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Raydugin, Yuri G. Modern Risk Quantification in Complex Projects. Oxford University Press, 2020. http://dx.doi.org/10.1093/oso/9780198844334.001.0001.

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There are multiple complaints that existing project risk quantification methods—both parametric and Monte Carlo—fail to produce accurate project duration and cost-risk contingencies in a majority of cases. It is shown that major components of project risk exposure—non-linear risk interactions—pertaining to complex projects are not taken into account. It is argued that a project system consists of two interacting subsystems: a project structure subsystem (PSS) and a project delivery subsystem (PDS). Any misalignments or imbalances between these two subsystems (PSS–PDS mismatches) are associated with the non-linear risk interactions. Principles of risk quantification are developed to take into account three types of non-linear risk interactions in complex projects: internal risk amplifications due to existing ‘chronic’ project system issues, knock-on interactions, and risk compounding. Modified bowtie diagrams for the three types of risk interactions are developed to identify and address interacting risks. A framework to visualize dynamic risk patterns in affinities of interacting risks is proposed. Required mathematical expressions and templates to factor relevant risk interactions to Monte Carlo models are developed. Business cases are discussed to demonstrate the power of the newly-developed non-linear Monte Carlo methodology (non-linear integrated schedule and cost risk analysis (N-SCRA)). A project system dynamics methodology based on rework cycles is adopted as a supporting risk quantification tool. Comparison of results yielded by the non-linear Monte Carlo and system dynamics models demonstrates a good alignment of the two methodologies. All developed Monte Carlo and system dynamics models are available on the book’s companion website.
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Book chapters on the topic "Multiplex amplification"

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El-Heliebi, Amin, Shukun Chen, and Thomas Kroneis. "Using Multiplex PCR for Assessing the Quality of Whole Genome Amplified DNA." In Whole Genome Amplification, 119–28. New York, NY: Springer New York, 2015. http://dx.doi.org/10.1007/978-1-4939-2990-0_9.

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Stiller, Mathias, and Tara L. Fulton. "Multiplex PCR Amplification of Ancient DNA." In Methods in Molecular Biology, 133–41. Totowa, NJ: Humana Press, 2011. http://dx.doi.org/10.1007/978-1-61779-516-9_17.

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Berbegall, Ana Pilar, Eva Villamón, Samuel Navarro, and Rosa Noguera. "Multiplex Ligation-dependent Probe Amplification (MLPA)." In Guidelines for Molecular Analysis in Archive Tissues, 215–24. Berlin, Heidelberg: Springer Berlin Heidelberg, 2011. http://dx.doi.org/10.1007/978-3-642-17890-0_33.

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Dash, Hirak Ranjan, Pankaj Shrivastava, and Surajit Das. "Amplification of Autosomal STR Markers by Multiplex PCR." In Springer Protocols Handbooks, 157–63. New York, NY: Springer US, 2020. http://dx.doi.org/10.1007/978-1-0716-0274-4_19.

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Moelans, Cathy B., Roel A. de Weger, and Paul J. van Diest. "Amplification Testing in Breast Cancer by Multiplex Ligation-Dependent Probe Amplification of Microdissected Tissue." In Methods in Molecular Biology, 107–18. Totowa, NJ: Humana Press, 2011. http://dx.doi.org/10.1007/978-1-61779-163-5_9.

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Moelans, Cathy B., Lilit Atanesyan, Suvi P. Savola, and Paul J. van Diest. "Methylation-Specific Multiplex Ligation-Dependent Probe Amplification (MS-MLPA)." In Methods in Molecular Biology, 537–49. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7481-8_27.

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Dash, Hirak Ranjan, Pankaj Shrivastava, and Surajit Das. "Amplification of Y Chromosome STR Markers by Multiplex PCR." In Springer Protocols Handbooks, 165–71. New York, NY: Springer US, 2020. http://dx.doi.org/10.1007/978-1-0716-0274-4_20.

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Dash, Hirak Ranjan, Pankaj Shrivastava, and Surajit Das. "Amplification of X Chromosome STR Markers by Multiplex PCR." In Springer Protocols Handbooks, 173–78. New York, NY: Springer US, 2020. http://dx.doi.org/10.1007/978-1-0716-0274-4_21.

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Kimpton, C., P. Gill, A. Urquhart, A. Walton, M. Adams, S. Watson, and E. Millican. "Automated DNA Profiling Employing Multiplex Amplification of Short Tandem Repeat Loci." In Advances in Forensic Haemogenetics, 309–11. Berlin, Heidelberg: Springer Berlin Heidelberg, 1994. http://dx.doi.org/10.1007/978-3-642-78782-9_80.

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Stirling, David. "Multiplex Amplification Refractory Mutation System for the Detection of Prothrombotic Polymorphisms." In PCR Protocols, 323–25. Totowa, NJ: Humana Press, 2003. http://dx.doi.org/10.1007/978-1-4612-0055-0_47.

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Conference papers on the topic "Multiplex amplification"

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Treerattrakoon, Kiatnida, Patraporn Luksirikul, Tararaj Dharakul, and Deanpen Japrung. "Isothermal amplification and graphene based detection of circulating multiplex miRNAs." In 2016 IEEE 16th International Conference on Nanotechnology (IEEE-NANO). IEEE, 2016. http://dx.doi.org/10.1109/nano.2016.7751320.

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van Slooten, H.-J., CC Kuijpers, CB Moelans, A. Horstman, S. Al-Janabi, JW Hinrichs, PJ van Diest, and M. Jiwa. "Abstract PD02-03: Added value of HER-2 amplification testing by multiplex ligation-dependent probe amplification (MLPA)." In Abstracts: Thirty-Fifth Annual CTRC‐AACR San Antonio Breast Cancer Symposium‐‐ Dec 4‐8, 2012; San Antonio, TX. American Association for Cancer Research, 2012. http://dx.doi.org/10.1158/0008-5472.sabcs12-pd02-03.

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Zhang, Ye, Pengfei Zhang, and Tza-Huei Wang. "Droplet-Based Digital Ratiometric Fluorescence Coding for Multiplex Nucleic Acid Amplification Testing." In 2019 IEEE 32nd International Conference on Micro Electro Mechanical Systems (MEMS). IEEE, 2019. http://dx.doi.org/10.1109/memsys.2019.8870782.

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KONWAR, K. M., I. I. MĂNDOIU, A. C. RUSSELL, and A. A. SHVARTSMAN. "IMPROVED ALGORITHMS FOR MULTIPLEX PCR PRIMER SET SELECTION WITH AMPLIFICATION LENGTH CONSTRAINTS." In 3rd Asia-Pacific Bioinformatics Conference. PUBLISHED BY IMPERIAL COLLEGE PRESS AND DISTRIBUTED BY WORLD SCIENTIFIC PUBLISHING CO., 2005. http://dx.doi.org/10.1142/9781860947322_0005.

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DORTA FERREIRA, ROBERTA, MARIA DE FATIMA SONATI, NATÁLIA DE OLIVEIRA MOTA, GISELE AUDREI PEDROSO, ELZA MIYUKI KIMURA BEZERRA, ANA SOLER, and JULIO DA LUZ. "Investigação de Deleções Talassêmicas Raras por Multiplex Ligation-dependent Probe Amplification (MLPA)." In XXIV Congresso de Iniciação Científica da UNICAMP - 2016. Campinas - SP, Brazil: Galoa, 2016. http://dx.doi.org/10.19146/pibic-2016-50648.

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Okino, Steven T., Michelle Kong, and Yan T. Wang. "Abstract 2213: Evaluation of bias associated with high-multiplex, target-specific pre-amplification." In Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1538-7445.am2015-2213.

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Peng, Quan, Ravi Vijaya Satya, Marcus Lewis, Pranay Randad, John DiCarlo, and Yexun Wang. "Abstract 4879: Reducing amplification artifacts in highly multiplex amplicon sequencing by using molecular barcodes." In Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1538-7445.am2015-4879.

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Isaksson, Magnus, Henrik Johansson, Fredrik Roos, Elin Falk, Lotte Moens, Olle Ericsson, and Mats Nilsson. "Abstract 1158: Single step multiplex amplification and barcoding of FFPE samples for Next-Gen sequencing." In Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC. American Association for Cancer Research, 2010. http://dx.doi.org/10.1158/1538-7445.am10-1158.

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Liu, Haiyan E., Melanie Triboulet, Amin Zia, Meghah Vuppalapaty, Evelyn Kidess-Sigal, John Coller, Vanita S. Natu, et al. "Abstract 1724: Genomic profiling of Vortex-enriched CTCs using whole genome amplification and multiplex PCR-based targeted next generation sequencing." In Proceedings: AACR Annual Meeting 2017; April 1-5, 2017; Washington, DC. American Association for Cancer Research, 2017. http://dx.doi.org/10.1158/1538-7445.am2017-1724.

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Rand, Casey M., Erin E. West, Min Yu, Debra E. Weese-Mayer, and Lawrence Jennings. "Multiplex Ligation-Dependent Probe Amplification To Detect Deletions And Duplications In The Ret Gene In Sudden Infant Death Syndrome Cases." In American Thoracic Society 2011 International Conference, May 13-18, 2011 • Denver Colorado. American Thoracic Society, 2011. http://dx.doi.org/10.1164/ajrccm-conference.2011.183.1_meetingabstracts.a3707.

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Reports on the topic "Multiplex amplification"

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Bilbiie, Florin, and Marc Melitz. Aggregate-Demand Amplification of Supply Disruptions: The Entry-Exit Multiplier. Cambridge, MA: National Bureau of Economic Research, December 2020. http://dx.doi.org/10.3386/w28258.

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Mears, C. A., S. E. Labov, and A. T. Barfknecht. Energy-resolving x-ray detectors with charge amplification due to multiple quasiparticle tunneling. Office of Scientific and Technical Information (OSTI), August 1993. http://dx.doi.org/10.2172/10184378.

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Lauring, Josh. Multiple Cooperating Oncogenes Drive Recurrent Breast Cancer-Associated Chromosomal Amplifications: Creation of Isogenic Human Cell Line Models. Fort Belvoir, VA: Defense Technical Information Center, July 2014. http://dx.doi.org/10.21236/ada612716.

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Lauring, Josh. Multiple Cooperating Oncogenes Drive Recurrent Breast Cancer-Associated Chromosomal Amplifications: Creation of Isogenic Human Cell Line Models. Fort Belvoir, VA: Defense Technical Information Center, July 2013. http://dx.doi.org/10.21236/ada583983.

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