Podcasts about rflp

In this episode, Dr. VanderWaal discusses the use of RFLP to identify and classify different PRRS virus strains and veterinarians' thoughts on that. Dr. VanderWall shares some results of a survey she conducted that shows most veterinarians don't think we should be using RFLPs anymore. But we don't have an alternative tool for that yet. What do you think of that?*Watch the full episode: https://www.swinecampus.com/blog-------------The Swine it Podcast Show is trusted and supported by innovative companies like:- Zinpro (https://zinpro.com/)- Gestal (http://jygatech.com/)- AB Vista (https://www.abvista.com/)- Adisseo (http://www.adisseo.com/)- Genesus (https://www.genesus.com/)- Evonik (https://animal-nutrition.evonik.com/en/species/swine/)- Cloudfarms (https://www.cloudfarms.com/)Give us a Rating & Review - http://getpodcast.reviews/id/1460280128

To track nearby viruses' lineages and update vaccination and biosecurity protocols, proper virus classification is necessary in the swine industry. To understand this better, Dr. VanderWaal explains what RFLP - restriction fragment length polymorphism - is and how we use it to identify different PRRS virus strains. We invite you to watch the complete episode!*Watch the full episode: https://www.swinecampus.com/blog-------------The Swine it Podcast Show is trusted and supported by innovative companies like:- Zinpro (https://zinpro.com/)- Gestal (http://jygatech.com/)- AB Vista (https://www.abvista.com/)- Adisseo (http://www.adisseo.com/)- Genesus (https://www.genesus.com/)- Evonik (https://animal-nutrition.evonik.com/en/species/swine/)- Cloudfarms (https://www.cloudfarms.com/)Give us a Rating & Review - http://getpodcast.reviews/id/1460280128

Dr. VanderWaal is an Associate Professor in the Department of Veterinary Population Medicine at the University of Minnesota. She shares her knowledge of how PRRS virus strains have been classified, especially focusing on the RFLP tool and what's changing in the way we classify those strains. Interesting, right?*Watch the full episode: https://www.swinecampus.com/blog-------------The Swine it Podcast Show is trusted and supported by innovative companies like:- Zinpro (https://zinpro.com/)- Gestal (http://jygatech.com/)- AB Vista (https://www.abvista.com/)- Adisseo (http://www.adisseo.com/)- Genesus (https://www.genesus.com/)- Evonik (https://animal-nutrition.evonik.com/en/species/swine/)- Cloudfarms (https://www.cloudfarms.com/)Give us a Rating & Review - http://getpodcast.reviews/id/1460280128

Recorded on location at the 2022 Conference on Crimes Against Women, each episode in this bonus series will deepen our understanding of DNA testing, how it supports cold case investigations and prosecutions, and what's ahead for crime solvers around the globe.DNA, FGG, ABO, RFLP, PCR, CODIS - what does it all mean? From the crime scene to the crime lab to the courtroom, DNA evidence collection, testing and results continue change the landscape of criminal investigations. We take a closer look at how DNA testing has evolved over the past century and the impact it has on investigations and securing convictions with a special focus on the successes of sexual assault kit testing. Patricia (Patti) Powers, JD, Attorney Advisor at AEquitas and Misty Mara Williamson, Forensic DNA Analyst III and DNA Laboratory Coordinator at the Marshall University Forensic Science Center, join the conversation as both experts in their fields and collaborators in bringing cold cases to justice. Patti and Misty co-presented, "Advancing Justice with DNA Technology", at the 2022 Conference on Crimes Against Women.

DNA paternity testing is the use of DNA profiles to determine whether an individual is the biological parent of another individual. Paternity testing can be especially important when the rights and duties of the father are in issue and a child's paternity is in doubt. Tests can also determine the likelihood of someone being a biological grandparent. Though genetic testing is the most reliable standard, older methods also exist, including ABO blood group typing, analysis of various other proteins and enzymes, or using human leukocyte antigen antigens. The current techniques for paternity testing are using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). Paternity testing can now also be performed while the woman is still pregnant from a blood draw. DNA testing is currently the most advanced and accurate technology to determine parentage. In a DNA paternity test, the result (called the 'probability of parentage) is 0% when the alleged parent is not biologically related to the child, and the probability of parentage is typically 99.99% when the alleged parent is biologically related to the child. However, while almost all individuals have a single and distinct set of genes, rare individuals, known as "chimeras", have at least two different sets of genes, which can result in a false negative result if their reproductive tissue has a different genetic make-up from the tissue sampled for the test. --- Send in a voice message: https://anchor.fm/law-school/message Support this podcast: https://anchor.fm/law-school/support

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.10.24.353425v1?rss=1 Authors: Kanyerezi, S., Nabisubi, P. Abstract: Introduction: Tuberculosis (TB) is the leading cause of morbidity and mortality globally, responsible for an estimated annual 10.0 million new cases and 1.3 million deaths among infectious diseases with Africa contributing a quarter of these cases in 2019. Classification of Mycobacterium tuberculosis (MTB) strains is important in understanding their geographical predominance and pathogenicity. Different studies have gone ahead to classify MTB using different methods. Some of these include; RFLP, spoligotyping, MIRU-VNTR and SNP set based phylogeny. The SNP set based classification has been found to be in concordance with the region of difference (RD) analysis of MTB complex classification system. In Uganda, the most common cause of pulmonary tuberculosis (PTB) is Uganda genotype of MTB and accounts for up to 70 % of isolates. Methods: Sequenced MTB genome samples were retrieved from NCBI and others from local sequencing projects. The genomes were subjected to snippy (a rapid haploid variant calling and core genome alignment) to call variants and annotate them. Outputs from snippy were used to classify the isolates into Uganda genotypes and Non Ugandan genotypes based on 62 SNP set. The Ugandan genotype isolates were later subjected to 413 SNP set and then to a pan genome wide association analysis. Results: 6 Uganda genotype isolates were found not to classify as either Uganda I or II genotypes based on the 62 SNP set. Using the 413 SNP set, the 6 Uganda genotype isolates were found to have only one SNP out of the 7 SNPs that classify the Uganda I genotypes. They were also found to have both missense and frameshift mutations within the ctpH gene whereas the rest of Uganda I that had a mutation within this gene, was a missense. Conclusion: Among the Uganda genotypes genomes, Uganda I genomes are unstable. We used publicly available datasets to perform analysis like mapping, variant calling, mixed infection, pan-genome analysis to investigate and compare evolution of the Ugandan genotype. Copy rights belong to original authors. Visit the link for more info

Join Michael Carnahan and Lisa O’Brien on Monday, March 4, 2019, at 8:00 p.m. Central, for our interview with Meghan Clement, MS, D-ABC, Forensic DNA and Serology Consultant with Clement Consulting, LLC.  We’ll talk about general DNA topics, including collection methods, equipment, testing methods, statistics and sources and types of DNA.  Ms. Clement began her career in law enforcement with the Albuquerque Police Department Crime Laboratory and she has held various director positions with Laboratory Corporation of American and was an integral member of the management team at Cellmark Forensics in Texas and maintained her technical expertise by performing technical reviews and case consultations.  She has testified over 370 times on cases that utilized all DNA technologies including RFLP, DQ Alpha/Polymarker, VNTR, STR, Y-STR and mtDNA. Ms. Clement holds a Bachelor’s degree in Biology from Westfield State College in Massachusetts and received her Master of Science in Forensic Science at the University of New Haven in West Haven, CT under the tutelage of Dr. Robert Gaensslen and Dr. Henry Lee.  We’re a live show and calls are always welcome at (347) 989-1171.     About Meghan Clement: https://clearandconvincingpodcast.wordpress.com/2019/03/01/season-2-episode-1-dna-information-with-meghan-clement-ms-d-abc/

For over fifty years, University of Utah has been a leader in human genetics, a field that is making precision medicine possible today. Ray Gesteland, Ph.D., professor emeritus in human genetics, describes the perfect storm of people and resources at the University of Utah that have since spurred discoveries of the genetic causes behind cystic fibrosis, colon cancer, and many more. In a second interview, Gesteland talks about the Alta Meeting and other events that led to the discovery of restriction fragment length polymorphism (RFLP), a tool that made mapping the human genome possible.

In 1980, a landmark paper described restriction fragment length polymorphism (RFLP), a method predateding high volume DNA sequencing, that could be used to identify disease-causing genes. Ray Gesteland, Ph.D., professor emeritus of human genetics at the University of Utah, remembers hearing about the technology from his colleagues as they were still working it out. He explains how the idea came about, what it is, and how it made modern genetics possible. In a second interview, Gesteland talks about what brings some of the world’s best geneticists to the University of Utah.

Due to increasing drug resistance, artemisinin-based combination chemotherapy (ACT) has become the first-line treatment of falciparum malaria in many endemic countries. Ethiopia has also adopted artemether/lumefantrine (AL) as first-line treatment in 2004 and its broad introduction was achieved in 2006. However, irreversible ototoxicity associated with AL has been reported and suggested to be a serious limitation in the use of ACT. The aim of this study was to compare ototoxicity, tolerability, and efficacy of ACT with that of quinine and atovaquone/proguanil in the treatment of uncomplicated falciparum malaria. 97 patients in Jimma area, Ethiopia with slide-confirmed malaria were randomly assigned to receive either artemether/lumefantrine or quinine or atovaquone/proguanil and followed-up on days 7, 28, and 90. Comprehensive audio-vestibular testing by pure tone audiometry (PTA), transitory evoked (TE) and distortion product (DP) otoacoustic emissions (OAE) and brain stem evoked response audiometry (BERA) was done before enrolment and on follow-up days. PTA and DP-OAE levels revealed transient significant cochlear hearing loss in patients treated with quinine but not in those treated with artemether/lumefantrine or atovaquone/proguanil. There was no evidence of drug-induced brain stem lesions by BERA measurements. Hence, there was no detrimental effect of a standard oral regimen of artemether/lumefantrine on peripheral hearing or brainstem auditory pathways in patients with uncomplicated falciparum malaria. In contrast, transient hearing loss was evidenced with quinine therapy due to temporary outer hair cell dysfunction. Reinfection and recrudescence were determined by RFLP of msp-1 and msp-2 genes. Mutations associated with drug resistance were characterized in Pfmdr1, Pfdhfr, Pfcytb, and Pfserca genes. Single nucleotide polymorphisms (SNPs) previously reported to be associated with resistance to the study drugs were identified in both recrudescent and treatment sensitive isolates. A total of seven recrudescences were obtained. The Pfmdr1 N86Y mutation was found in 84.5% of isolates. The triple mutation 51I, 59R, 108N of the Pfdhfr gene occurred in high frequency (83.3%) but no Pfcytb mutation was detected. Sequencing showed a variety of previously described and new mutations in the Pfserca gene. The prevalence of high degree of mutations in Pfmdr1 and Pfdhfr is a reminiscent of the impact of previously used drugs in the area, chloroquine and sulfadoxine/pyrimethamine as first-line treatments. The broad introduction of AL and the cessation of the former drug regimens might probably change the current distribution of polymorphisms, possibly leading to decreased sensitivity to AL in the future. Continuous surveillance of molecular patterns in this region is, therefore, recommended.

Background: The emergence of drug resistance is a major problem in malaria control. Combination of molecular genotyping and characterization of mutations or single nucleotide polymorphisms ( SNPs) correlated with drug resistance can provide information for subsequent surveillance of existing and developing drug resistance patterns. The introduction of artemether/lumefantrine (AL) as first-line treatment, never used before in Ethiopia, allowed the collection of baseline data of molecular polymorphisms before a selection due to AL could occur. Method: 97 patients with uncomplicated falciparum malaria were recruited from April to June 2006 and treated with either AL, quinine (Q) or atovaquone/proguanil (AP) in Jimma University Hospital, Ethiopia. Mutations or SNPs associated with resistance to these drugs were analysed by RFLP (pfdhfr, pfmdr1) and sequencing of the target genes (pfcytb, pfserca). Results: SNPs previously reported to be associated with resistance to the study drugs were identified in recrudescent and treatment sensitive isolates. A total of seven recrudescences were obtained. The pfmdr1 N86Y mutation was found in 84.5% of isolates. The triple mutation 51I,59R,108N of the pfdhfr gene occured in high frequency (83.3%) but no pfcytb mutation was detected. Sequencing showed a variety of previously described and new mutations in the pfserca gene. Conclusion: The prevalence of mutations was in accordance with the expected patterns considering recent drug regimens. The broad introduction of AL and the cessation of former drug regimens might probably change the current distribution of polymorphisms, possibly leading to decreased sensitivity to AL in future. Continuous surveillance of molecular patterns in this region is, therefore, recommended.

Background: Gastric inhibitory polypeptide ( GIP) is postulated to be involved in type 2 diabetes mellitus and obesity. It exerts its function through its receptor, GIPR. We genotyped three GIPR SNPs (rs8111428, rs2302382 and rs1800437) in German families with at least one obese index patient, two case-control studies and two cross-sectional population-based studies. Methods: Genotyping was performed by MALDI-TOF, ARMS-PCR and RFLP. The family-study: 761 German families with at least one extremely obese child or adolescent (n = 1,041) and both parents ( n = 1,522). Case-control study: ( a) German obese children ( n = 333) and (b) obese adults ( n = 987) in comparison to 588 adult lean controls. The two cross-sectional population-based studies: KORA ( n = 8,269) and SHIP ( n = 4,310). Results: We detected over-transmission of the A-allele of rs2302382 in the German families (pTDT-Test = 0.0089). In the combined case-control sample, we estimated an odd ratio of 1.54 (95% CI 1.09; 2.19, pCA-Test = 0.014) for homozygotes of the rs2302382 A-allele compared to individuals with no A-allele. A similar trend was found in KORA where the rs2302382 A-allele led to an increase of 0.12 BMI units (p = 0.136). In SHIP, however, the A-allele of rs2302382 was estimated to contribute an average decrease of 0.27 BMI units (p-value = 0.031). Conclusion: Our data suggest a potential relevance of GIPR variants for obesity. However, additional studies are warranted in light of the conflicting results obtained in one of the two population-based studies.

625 adult unfed I. ricinus ticks from three recreational areas located near Munich and Passau, 275 adult engorged I. ricinus ticks from dogs of Bavaria and Baden-Württemberg and 25 adult engorged I. ricinus ticks from cattle of an area in Switzerland, which is endemic for bovine granulocytic ehrlichiosis, were collected over a specific period (3/2003-3/2004). The ticks were examined for an infection with B. burgdorferi sensu lato spp., A. phagocytophilum and piroplasms by the use of Real-Time PCR and nested PCR. In addition RFLP analysis and sequencing were chosen for the differentiation of the species and OspA types of B. burgdorferi sensu lato. The examination of unfed ticks resulted in a prevalence of 35,4% for B. burgdorferi sensu lato, 4,5% for A. phagocytophilum and 1,3% for piroplasms. There was no significant difference for the infection rates of B. burgdorferi sensu lato between the different sampling areas, whereas A. phagocytophilum showed a significantly higher prevalence in one sampling side in Munich and a significantly lower prevalence in Passau. Apart from infections with only one pathogen, coinfections with B. burgdorferi sensu lato and A. phagocytophilum could be detected in 1,1% of the unfed ticks, with a local cluster in one area in Munich and 0,3% of the unfed ticks showed a coinfection with B. burgdorferi sensu lato and piroplasms. Prevalence rates of 8,4%, 4,7% and 5,1% were identified for B. burgdorferi sensu lato, A. phagocytophilum and piroplasms respectively in engorged ticks from dogs, 0,7% of these ticks were coinfected with B. burgdorferi sensu lato and A. phagocytophilum. The examination of engorged ticks from cattle revealed in a prevalence rate of 8,0% for B. burgdorferi sensu lato and 60,0% for A. phagocytophilum. The high infection rate of A. phagocytophilum probably resulted from an infection of the cattle with this pathogen. The difference in the prevalence rate of B. burgdorferi sensu lato in unfed and engorged ticks might be caused by their distinct geographical origin, the degree of blood uptake and different factors in the blood of the different host species which are able to protect the host from being infected with Borrelia. The differentiation of B. burgdorferi sensu lato into the species and OspA types showed that the clinically relevant species B. afzelii, B. burgdorferi sensu stricto and B. garinii, in which B. garinii was represented by the OspA types 3, 4, 5, 6 and 8 could be detected. Additionally, B. valaisiana, a species which is suspected of being pathogenic to humans and the recently described new Borrelia genospecies, B. spielmanii (previously A14S), were detected. Alltogether a broad heterogeneity for Borrelia species and subspecies (classified by OspA types) could be observed in unfed ticks, above all in one sampling side in Munich. Ticks engorgd from dogs and cattle showed a less heterogeneous pattern of distribution.

Die Grundlage für die Entstehung von Tumoren bildet der Erwerb genetischer Veränderungen, wobei neben Mutationen wie Chromosomeninstabilitäten, Insertionen, Deletionen oder Basenmutationen auch epigenetische Vorgänge von Bedeutung sind. Hierzu zählt die veränderte Methylierung von DNA in neoplastisch transformierten Zellen, die bei einigen Genen in deren transkriptionellen Inaktivierung resultiert. Im Rahmen der vorliegenden Arbeit sollten Kolon- und Magenkarzinome auf bestehende Methylierungsdefekte in zwei Genen untersucht werden. Im ersten Teil wurde ein in die Mismatch-Reparatur involviertes Gen, das hMLH1-Gen, untersucht. Gastrointestinale Karzinome wurden hierbei hinsichtlich eines Zusammenhangs zwischen ihrer Mikrosatelliten- instabilität und einer möglichen Methylierung von Cytosinbasen im hMLH1-Promotor analysiert. Hierfür kam die Bisulfitmethode zur Anwendung, bei der die Umwandlung von unmethylierten Cytosinbasen zu Uracil erfolgt und die damit die Grundlage für die anschließende methylierungssensitive PCR bildet. Insgesamt ließ sich die Mikrosatelliten- instabilität bei sieben von neun untersuchten Karzinomproben auf Methylierung der proximalen Promotorsequenz zurückführen. Der Methylierungsstatus der beiden übrigen mikrosatelliteninstabilen Tumoren konnte nicht eindeutig bestimmt werden, obwohl die Vollständigkeit der Bisulfitmodifikation über den Nachweis des maternalen Methylierungsmusters des Exons 1 von SNRPN bestätigt wurde. Überdies konnte eine Deletion dieser Sequenz mittels Überprüfung der unbehandelten DNA ausgeschlossen werden. Gegenstand des zweiten Teils dieser Arbeit war die Untersuchung des CD95-Gens, eines Mitglieds der Tumor- Nekrose-Faktor-Rezeptor-Familie (sog. Todesrezeptoren), das eine entscheidende Rolle bei der ungestörten Apoptose von Zellen spielt. Grundlage hierfür waren die Ergebnisse von Peli et al., 1999, denen zufolge die CD95-Expression in mehreren etablierten Säugetierzellinien durch onkogenes Ras (H-Ras) herunterreguliert wird, wodurch sich eine Resistenz der Zellen für CD95L-induzierte Apoptose entwickelt. Hinweise aus den genannten Untersuchungen, die Inaktivierung von CD95 sei möglicherweise durch Methylierung des Promotors verursacht, sollte in der vorliegenden Arbeit anhand von gastrointestinalen Karzinomen näher analysiert werden. Hierzu wurde der Methylierungsstatus von Promotor, Exon 1 und Intron-Enhancer des CD95-Gens von 17 Tumorproben mit Ras-Mutationen (KRAS2-Mutation in Kodon 12) mittels methylierungssensitiver Restriktionsenzyme, nachfolgendem Southernblot und Hybridisierung ermittelt. Bei 16 Tumoren erbrachte die Restriktionsanalyse für Sma I ein Restriktionsmuster, das eine Methylierung der Cytosine der CCCGGG-Motive im genannten Bereich ausschließt. Die nicht geschnittene Sma I-Sequenz in einer Tumor-DNA (Position 143901) wurde auch für die Normal-DNA nachgewiesen und erwies sich als RFLP. Ein aufgrund dieses Ergebnisses postulierter Polymorphismus innerhalb einer Sma I- Schnittstelle konnte durch Sequenzierung der isolierten Sequenz aus dem entsprechenden Tumor verifiziert werden. Möglicherweise ist die Entstehung dieses Polymorphismus zwar die Folge einer ehemaligen Methylierung des Cytosins in der Konstellation CpG; allerdings konnte nachgewiesen werden, daß der entsprechende Nukleotidaustausch nicht im Rahmen der Karzinogenese erfolgt war, da die DNA aus PBL des gleichen Patienten die identische Sequenz aufwies. Insgesamt wurde somit für keinen der untersuchten Tumoren eine Abschaltung der CD95-Expression aufgrund einer Methylierung des Gens durch aktiviertes, onkogenes Ras festgestellt.

N-Acylhomoserinlactones (AHL) are signalling molecules in gram-negative bacteria, which regulate, in a cell density dependent way, important interactive functions. This phenomenon is known as quorum-sensing. This work characterised the microbial ecology of the autoinducer (AHL) producing bacteria Serratia liquefaciens MG1 and Pseudomonas putida IsoF in the rhizosphere of tomato plants. Gfp- and rfp-tagged strains of the AHL producing wiltypes S. liquefaciens MG1 and P. putida IsoF were compared with its AHL-negative mutants, which were unable to produce AHL. Two kinds of plants cultivating systems were used: a defined axenic system and a complex soil system. The characterisation of the root colonisation behaviour was performed using confocal laserscanning microscopy (CLSM) and cell counting of bacteria. Fluorescence in situ hybridisation (FISH) and terminal restrictions fragment length polymorphism (t-RFLP)-techniques were used to examinate shifts of the bacterial population in the rhizosphere on tomato plants. The effective in situ production and spreading of AHL on tomato roots was demonstrated with P. putida IsoF using an AHL-sensor strain P. putida F117 pKR-C12. AHL was produced in effective concentrations in the rhizosphere of tomato plants and influenced the bacterial rhizosphere population. However, the AHL-production had no influence on the colonization behaviour of the AHL-producing strains S. liquefaciens MG1 and P. putida IsoF.

Im Literaturüberblick der vorliegenden Arbeit wird besonders auf die ökonomische und ökologische Bedeutung der Flusskrebse, den derzeitigen Stand der Forschung die Krebspest betreffend und die aktuelle taxonomische Klassifikation des Krebspesterregers, Aphanomyces astaci, eingegangen. Ziel der vorliegenden Arbeit war es, eine zuverlässige und schnelle Methode zum spezifischen und sensitiven Nachweis von Aphanomyces astaci direkt aus Krebsgewebe zu entwickeln. Zu diesem Zweck wurde zunächst ein kommerzielles Fertigkit (DNeasy® Tissue Kit, Qiagen) als geeignete DNA-Extraktionsmethode ausgewählt. Als Untersuchungsmaterial wurde die Abdominalkutikula von Edelkrebsen (Astacus astacus) eingesetzt. Anhand von Genomanalysen der ITS-Regionen (Genabschnitte, die zwischen den die ribosomale RNA kodierenden Genen liegen) verschiedener Oomycota wurden Oligonukleotid-Primer konstruiert, optimiert und auf ihre Eignung überprüft. Ausgehend von diesen Untersuchungen wurden zwei PCR-Protokolle etabliert: eine Semi-Nested und eine Nested PCR. Die Semi-Nested PCR, die sich für die Routinediagnostik von Aphanomyces astaci als wenig geeignet erwiesen hat, bietet das Potential, als Grundlage für weiterführende Studien an anderen Arten der Gattung Aphanomyces zu dienen. Die Nested PCR mit den äußeren Primern NS 166/NS 681 und den inneren Primern BO 525/BO 640 ergab mit allen 20 untersuchten Aphanomyces astaci-Stämmen ein PCR-Produkt von etwa 115 bp, das mit einer anschließenden Restriktionsenzym-Analyse mit der Endonuklease Hph I verifiziert werden konnte. Die Nested PCR zeigte sich hochspezifisch gegenüber Aphanomyces astaci, während die DNA von anderen bei Krebsen und im Wasser vorkommenden Krankheitserregern und die DNA von Wirtsgewebe nicht amplifiziert wurde, was Grundvoraussetzung für den Einsatz dieser PCR als Standardmethode ist. Die Nachweisgrenze der Nested PCR lag bei Verwendung von Plasmid-DNA bei 1,9 genomischen Einheiten und bei Einsatz von genomischer DNA aus Pilzmyzel bei 1 ag. Als Zusammenfassung 172 Aphanomyces astaci-Sporen in die DNA-Extraktion eingesetzt wurden, lag die Nachweisgrenze bei 1 Spore. Zusätzlich wurden Infektionsversuche durchgeführt, bei denen Edelkrebse mit 10, 100 und 1000 Aphanomyces astaci-Zoosporen/ml infiziert wurden. Ein positives PCR-Ergebnis konnte ab Tag 2 post expositionem (entspricht dem Tag der ersten Probenentnahme) mit der beschriebenen Methode bei allen untersuchten Tieren beobachtet werden. Schließlich wurde eine Validierung der entwickelten Nested PCR unter Einsatz von Wirtskutikula anhand 19 diagnostischer Proben unterschiedlicher Herkunft innerhalb Europas (aus Deutschland, der Schweiz, Österreich, Schweden und England) durchgeführt und die Ergebnisse mittels RFLP verifiziert. Mit der in der vorliegenden Arbeit entwickelten molekularbiologischen Nachweismethode steht ein Verfahren zur Verfügung, das eine zuverlässige Diagnose von Aphanomyces astaci bereits im Anfangsstadium der Infektion direkt aus Krebsgewebe ermöglicht und innerhalb von 12 bis 15 Stunden (Sektion, DNA-Extraktion, Nested PCR, Agarosegel-Elektrophorese und bestätigende Restriktionsenzym-Analyse) durchführbar ist. Diese Methode ist somit ein geeignetes und zuverlässiges Mittel, um in Programmen zur Seuchenbekämpfung der Krebspest (Aphanomyces astaci) eingesetzt zu werden.

We investigated the Nco I restriction fragment length polymorphism (RFLP) of the tumor necrosis factor beta (TNFB) gene in 173 patients with systemic lupus erythematosus (SLE), 192 unrelated healthy controls, and eleven panel families, all of German origin. The phenotype frequency of the TNFB*I allele was significantly increased in patients compared to controls (63.6% vs 47.1%, RR = 1.96, p