An assessment of trap efficiency to estimate coho salmon smolt abundance in a small Alaskan stream

Thesis (M.S.) University of Alaska Fairbanks, 2004Smolt abundance is commonly estimated using trap efficiency-based methods; however, few studies have investigated the accuracy of trap efficiency estimates. The objectives of this study were to: (1) test the hypotheses that (i) trap efficiency is not affected by release timing nor release distance, (ii) trap efficiency-based estimates of smolt abundance are concordant with smolt-adult mark-recapture estimates, and (2) evaluate if water level and turbidity influence trap efficiency. In Deep Creek, Alaska, during 2001 and 2002, coho salmon Oncorhynchus kisutch smolt abundance was estimated using trap efficiency-based methods and compared to independent smolt-adult mark-recapture estimates. Marked smolts were released at two times of day (1200 hours and 0000 hours) and two release distances upstream of the trap (400 m and 1500 m) every 2 to 4 d throughout each year. Trap efficiency estimates were highly variable (range 0%-55%) and trap efficiency-based estimates of abundance were not concordant with smolt-adult mark-recapture estimates. Release timing and turbidity significantly influenced trap efficiency, whereas release distance did not. Several assumptions of the trap efficiency approach were not met, which produced biased estimates and conflicting results among years when comparing estimation techniques. These results suggest that assumptions of the trap efficiency-based methods be fully assessed to accurately estimate smolt abundance

Controlled Disassembly and Purification of Functional Viral Subassemblies Using Asymmetrical Flow Field-Flow Fractionation (AF4)

Viruses protect their genomes by enclosing them into protein capsids that sometimes contain lipid bilayers that either reside above or below the protein layer. Controlled dissociation of virions provides important information on virion composition, interactions, and stoichiometry of virion components, as well as their possible role in virus life cycles. Dissociation of viruses can be achieved by using various chemicals, enzymatic treatments, and incubation conditions. Asymmetrical flow field-flow fractionation (AF4) is a gentle method where the separation is based on size. Here, we applied AF4 for controlled dissociation of enveloped bacteriophage phi 6. Our results indicate that AF4 can be used to assay the efficiency of the dissociation process and to purify functional subviral particles.Peer reviewe

Asymmetrical Flow Field-Flow Fractionation on Virus and Virus-Like Particle Applications

Asymmetrical flow field-flow fractionation (AF4) separates sample components based on their sizes in the absence of a stationary phase. It is well suited for high molecular weight samples such as virus-sized particles. The AF4 experiment can potentially separate molecules within a broad size range (~103−109 Da; particle diameter from 2 nm to 0.5−1 μm). When coupled to light scattering detectors, it enables rapid assays on the size, size distribution, degradation, and aggregation of the studied particle populations. Thus, it can be used to study the quality of purified viruses and virus-like particles. In addition to being an advanced analytical characterization technique, AF4 can be used in a semi-preparative mode. Here, we summarize and provide examples on the steps that need optimization for obtaining good separation with the focus on virus-sized particles

Asymmetrical Flow Field-Flow Fractionation on Virus and Virus-Like Particle Applications

Asymmetrical flow field-flow fractionation (AF4) separates sample components based on their sizes in the absence of a stationary phase. It is well suited for high molecular weight samples such as virus-sized particles. The AF4 experiment can potentially separate molecules within a broad size range (~103−109 Da; particle diameter from 2 nm to 0.5−1 μm). When coupled to light scattering detectors, it enables rapid assays on the size, size distribution, degradation, and aggregation of the studied particle populations. Thus, it can be used to study the quality of purified viruses and virus-like particles. In addition to being an advanced analytical characterization technique, AF4 can be used in a semi-preparative mode. Here, we summarize and provide examples on the steps that need optimization for obtaining good separation with the focus on virus-sized particles

Avkorporatiseringen i Finland, En studie av FFC i det statliga kommittéväsendet 1981-1998

Only abstract. Paper copies of master’s theses are listed in the Helka database (http://www.helsinki.fi/helka). Electronic copies of master’s theses are either available as open access or only on thesis terminals in the Helsinki University Library.Vain tiivistelmä. Sidottujen gradujen saatavuuden voit tarkistaa Helka-tietokannasta (http://www.helsinki.fi/helka). Digitaaliset gradut voivat olla luettavissa avoimesti verkossa tai rajoitetusti kirjaston opinnäytekioskeilla.Endast sammandrag. Inbundna avhandlingar kan sökas i Helka-databasen (http://www.helsinki.fi/helka). Elektroniska kopior av avhandlingar finns antingen öppet på nätet eller endast tillgängliga i bibliotekets avhandlingsterminaler.Undersökningens syfte är att utreda en eventuell avkorporatisering i Finland, genom att studera Finlands Fackförbunds Centralorganisation i det statliga kommittéväsendet under åren 1981-1998. Avsikten är då att granska huruvida denna traditionella korporatistiska kanal har försvagats. FFC är Finlands största intresseorganisation, och därför är det intressant att se om denna, åtminstone tidigare så inflytelserika organisation, har mistat en del av sin makt, eller om den har hittat nya kanaler för påtryckningsverksamhet. Därtill diskuteras därför eventuella nya arbetsmetoder, och framför allt lobbyism. Som avslutning diskuteras också hur Finlands inträde i Europeiska Unionen 1995 har flyttat över en del av beslutsprocessen till ett övernationellt plan, och vilka intresseorganisationernas möjligheter är att utöva påtryckningsverksamhet gällande EU-ärenden i Finland, samt ärenden som besluts inom de europeiska institutionerna. Undersökningens baserar sig delvis på Leif Lewins teori om det ”lossnande greppet”, och delvis på Munk Christiansen & Rommetvedts studier av avkorporatiseringen i Norge och i Danmark. Lewins klassificering av intresseorganisationerna på basen av deras storlek, och dessas effekter på samhället i termer av konflikter eller goda relationer ligger som grund för diskussionen krings FFCs ställning idag. Därefter diskuteras Munk Christiansens & Rommetvedts teorier i ljuset av undersökningen av FFC, och den eventuellt nya arbetsmetoden, lobbyism. Studien av FFC i det statliga kommittéväsendet 1981-1998 visar en tydlig, och knappast särskilt överraskande, utveckling. De organiserade intressena, och i det här fallet FFC, har mistat en del av sitt tidigare så stora inflytande i inom kommittéväsendet. Orsakerna till denna utveckling, och slutsaterna man kan dra därav, är dock inte lika entydiga. Utvecklingen inom den korporatistiska kanalen för påtryckningsverksamhet är relaterad till förändringar inom de politiska såväl som de administrativa institutionerna och till samhälleliga strukturförändringar på det sociala och det ekonomiska planet. Då antalet kommittéer under senare tid har minskat märkbart, har naturligtvis också tillträdet till den här arenan för påtrycknngsverksamhet begränsats. Minskningen av antalet kommittéer över lag beror i sin tur på färre samhälleliga strukturförändringar, samt på att man flyttat över en del av kommittéernas arbete till andra organ, både på lokal och nationell nivå, samt på övernationell nivå. Däremot innebär detta inte nödvändigtvis att FFCs inflytande skulle ha minskat, då de allt mera har gått över till andra, idag mera effektiva arbetsmetoder som till exempel till lobbyism

Ribosome profiles and riboproteomes of healthy and Potato virus A- and Agrobacterium-infected Nicotiana benthamiana plants

Nicotiana benthamiana is an important model plant for plant-microbe interaction studies. Here, we compared ribosome profiles and riboproteomes of healthy and infected N. benthamiana plants. We affinity purified ribosomes from transgenic leaves expressing a FLAG-tagged ribosomal large subunit protein RPL18B of Arabidopsis thaliana. Purifications were prepared from healthy plants and plants that had been infiltrated with Agrobacterium tumefaciens carrying infectious cDNA of Potato virus A (PVA) or firefly luciferase gene, referred to here as PVA- or Agrobacterium-infected plants, respectively. Plants encode a number of paralogous ribosomal proteins (r-proteins). The N. benthamiana riboproteome revealed approximately 6600 r-protein hits representing 424 distinct r-proteins that were members of 71 of the expected 81 r-protein families. Data are available via ProteomeXchange with identifier PXD011602. The data indicated that N. benthamiana ribosomes are heterogeneous in their r-protein composition. In PVA-infected plants, the number of identified r-protein paralogues was lower than in Agrobacterium-infected or healthy plants. A. tumefaciens proteins did not associate with ribosomes, whereas ribosomes from PVA-infected plants co-purified with viral cylindrical inclusion protein and helper component proteinase, reinforcing their possible role in protein synthesis during virus infection. In addition, viral NIa protease-VPg, RNA polymerase NIb and coat protein were occasionally detected. Infection did not affect the proportions of ribosomal subunits or the monosome to polysome ratio, suggesting that no overall alteration in translational activity took place on infection with these pathogens. The riboproteomic data of healthy and pathogen-infected N. benthamiana will be useful for studies on the specific use of r-protein paralogues to control translation in infected plants.Peer reviewe

Native RNA purification method for small RNA molecules based on asymmetrical flow field-flow fractionation

RNA molecules provide promising new possibilities for the prevention and treatment of viral infections and diseases. The rapid development of RNA biology and medicine requires advanced methods for the purification of RNA molecules, which allow fast and efficient RNA processing, preferably under non-denaturing conditions. Asymmetrical flow field-flow fractionation (AF4) enables gentle separation and purification of macromolecules based on their diffusion coefficients. The aim of the study was to develop an AF4 method for efficient purification of enzymatically produced antiviral small interfering (si)RNA molecules and to evaluate the overall potential of AF4 in the separation of short single-stranded (ss) and double-stranded (ds) RNA molecules. We show that AF4 separates monomeric ssRNA from dsRNA molecules of the same size and monomeric ssRNA from multimeric forms of the same ssRNA. The developed AF4 method enabled the separation of enzymatically produced 27-nt siRNAs from partially digested substrate dsRNA, which is potentially toxic for mammalian cells. The recovery of AF4-purified enzymatically produced siRNA molecules was about 70%, which is about 20% higher than obtained using anion-exchange chromatography. The AF4-purified siRNAs were not toxic for mammalian cells and fully retained their biological activity as confirmed by efficient inhibition of herpes simplex virus 1 replication in cell culture. Our work is the first to develop AF4 methods for the separation of short RNA molecules.Peer reviewe

Analysis and purification of ssRNA and dsRNA molecules using asymmetrical flow field flow fractionation

Robust RNA purification and analysis methods are required to support the development of RNA vaccines and therapeutics as well as RNA interference-based crop protection solutions. Asymmetrical flow field -flow fractionation (AF4) is a gentle native purification method that applies liquid flows to separate sample components based on their hydrodynamic sizes. We recently showed that AF4 can be utilized to separate RNA molecules that are shorter than 110 nucleotides (nt), but the performance of AF4 in the analysis and purification of longer RNA molecules has not been previously evaluated. Here, we studied the perfor-mance of AF4 in separation of single-stranded (ss) and double-stranded (ds) RNA molecules in the size range of 75-6400 nt. In addition, we evaluated the power of AF4 coupling to different detectors, allow-ing separation to be combined with data collection on yield as well as molecular weight ( MW ) and size distribution. We show that AF4 method is applicable in RNA purification, quality control, and analytics, and results in good recoveries of ssRNA and dsRNA molecules. In addition, our results demonstrate the utility of AF4 multidetection platforms to study biophysical properties of long RNA molecules.(c) 2022 The Author(s). Published by Elsevier B.V.This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ )Peer reviewe

Native RNA Purification Method for Small RNA Molecules Based on Asymmetrical Flow Field-Flow Fractionation

RNA molecules provide promising new possibilities for the prevention and treatment of viral infections and diseases. The rapid development of RNA biology and medicine requires advanced methods for the purification of RNA molecules, which allow fast and efficient RNA processing, preferably under non-denaturing conditions. Asymmetrical flow field-flow fractionation (AF4) enables gentle separation and purification of macromolecules based on their diffusion coefficients. The aim of the study was to develop an AF4 method for efficient purification of enzymatically produced antiviral small interfering (si)RNA molecules and to evaluate the overall potential of AF4 in the separation of short single-stranded (ss) and double-stranded (ds) RNA molecules. We show that AF4 separates monomeric ssRNA from dsRNA molecules of the same size and monomeric ssRNA from multimeric forms of the same ssRNA. The developed AF4 method enabled the separation of enzymatically produced 27-nt siRNAs from partially digested substrate dsRNA, which is potentially toxic for mammalian cells. The recovery of AF4-purified enzymatically produced siRNA molecules was about 70%, which is about 20% higher than obtained using anion-exchange chromatography. The AF4-purified siRNAs were not toxic for mammalian cells and fully retained their biological activity as confirmed by efficient inhibition of herpes simplex virus 1 replication in cell culture. Our work is the first to develop AF4 methods for the separation of short RNA molecules