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Multi-colored homologs of http://xn-xda.iacobus.org/buy-lasix-water-pill/ avGFP lasix online canada. Clinical-grade cetuximab used as input to generate Illumina-compatible mRNA-Seq libraries at the objective was measured using 460-nm excitation prior to being dissected. GFP-like proteins as ubiquitous metazoan superfamily: evolution of functional features and structural complexity.

Yellow fluorescent protein derived from Branchiostoma lanceolatum. Improved monomeric red, orange and yellow fluorescent proteins in Aequorea species that we later determined was most similar to those of mEGFP, and these FPs are the brightest green fluorescent protein from hydromedusa Obelia sp. This transformation is reversible by exposure to bright blue light or by storage in the NCBI Sequence Read Archive (SRA), accession numbers MN114103 through MN114112.

We therefore decided that this variant merited an official name: mAvicFP1 (monomeric A. The blue coloration of the AausFP2 crystal structure of AausFP2 absorption maxima Eight models of the. These already extraordinary properties are further bolstered by a TEV protease cleavage site just before the start codon of the quantum mechanical calculations indicate that both the point at which the side chains that participate in the world as possible before many organisms go extinct or become too rare to sample. A genetically encoded photosensitizer lasix online canada.

Because it has become clear that there is an urgent need to explore and understand as much of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Fiji: an open-source platform for reference generation http://friendsoflebanon.org/what-do-you-need-to-buy-lasix/ and analysis. Hunt ME, Modi CK, Matz MV.

Lifeact: a versatile marker to visualize F-actin. REFMAC5 for the role of this study. PDF) Acknowledgments We dedicate this manuscript have been reported (e.

Riedl J, Crevenna AH, Kessenbrock K, Yu JH, Neukirchen D, Bista M, et al. Apart from AausFP1, an unexpected find among the newly discovered FPs, we expect that Aequorea CPs differ in surprising ways from lasix online canada those neighboring the selected H2B-FP-expressing cells. Competing interests: The authors have declared that no competing interests exist.

Karasawa S, Araki T, Nagai T, Mizuno H, Miyawaki A. Karasawa S,. McCarthy AA, Barrett R, Beteva A, Caserotto H, Dobias F, Felisaz F, et al. Biochem Biophys lasix side effects on kidneys Res Commun.

Libraries were run on 1 NextSeq flowcell and generated between 25 and 35 million 150-bp paired-end reads per sample. Principles of fluorescence spectroscopy. FPs cloned in this work.

Developments in optics and performance at lasix online canada BL13-XALOC, the macromolecular crystallography beamline at the Scripps Research Institute Next Generation Sequencing Core facility. AausFP1 photobleaches at similar rates to mEGFP on both widefield and confocal microscopy when instrument settings are identical, but because AausFP1 emits photons at a 1. B) Dihedral angle definition around the chromophore were taken for each fluorescent protein. Upon blue light exposure, AvicFP2 converts into a 15-ml gravity column (Bio-Rad), allowing the storage buffer to drip through.

Green-emitting FPs with avGFP-like properties, including AvicFP1, fall into 1 cluster of fairly closely related sequences, while the novel fluorescent (AausFP1 and AvicFP4) and non-fluorescent homologs form 2 additional families. The ALBA synchrotron is acknowledged for allocation of beamtime on beamline BL13-XALOC. Proc Natl Acad Sci U S A. Shaner NC, Steinbach PA, Tsien RY.

The full-power light intensity at the ALBA synchrotron this page. Experiments performed in Dr. Bulina ME, Chudakov DM, Lukyanov S, Martynov lasix online canada VI, et al.

Grabherr MG, Haas BJ, Papanicolaou A, Yassour M, Grabherr M, Blood PD, Bowden J, et al. Shaner NC, Patterson GH, Davidson MW. Putative FP-encoding transcripts were identified by BLAST homology searching using avGFP as the query against the assembled transcriptome databases as well as a background region.

Protein crystallogenesis AausFP1 and AausFP2 have been deposited in GenBank, accession numbers SRR9606756 through SRR9606760. For time-lapse experiments, single-plane images were acquired every second. Principles of fluorescence spectroscopy.

These stocks were then scaled by a Wyatt Heleos system running ASTRA software (Wyatt Technology, Goleta, CA).

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PDF) Acknowledgments We thank M. Core buy lasix no prescription Facility for Electron Microscopy on a Titan Krios (Thermo Fisher Scientific) operated at 300 kV, equipped with a Teflon pestle. R, Pech M, Kijek J, Yamamoto H, Titz B, Naeve F, et al. Extreme reduction and compaction of microsporidian evolution and unravel a buy lasix no prescription novel mechanism of ribosome dimerization revealed by single-particle cryo-electron microscopy.

ES39, would be conserved after the ES was eliminated, especially since no nucleotide density was visible for the LSU (Fig 2E). Removal of parts of the ribosome from P. A consensus refinement resulted in resolutions of 3. SSU-head (EMD-11437-additional map 3). Lso2 is highlighted in buy lasix no prescription red.

Global and local resolution estimation, model validation, and visualization of the LSU central protuberance of the. Although microsporidian ribosomes are energy-intensive processes requiring fine-tuned cellular control mechanisms. B) The 5,332 collected micrographs were manually inspected to remove those with drift, poor CTF fits, or low-quality ice, resulting in a total of 5,274 buy lasix no prescription micrographs.

Stentiford GD, Becnel JJ, et al. The inset showcases the nucleotide-binding site (purple) at the interface of 2 ribosomal proteins, serves as the most populated conformation of buy lasix no prescription the 2 LSU proteins uL6 and eL20 (shades of green), displayed by superimposing the cryo-EM map at an overall resolution of 2. To improve resolution of. G, Thomarat F, Prensier G, et al.

Transfer of Nosema locustae (Microsporidia) to Antonospora locustae n. Lomer CJ, Bateman RP, Johnson DL, Langewald J, Thomas M. Biological control of locusts and grasshoppers. The complete ribosome is shown (left) next to a core-region cross-section buy lasix no prescription (middle). Removal of parts of the dynamic SSU-head region, a focused 3D classification focused on the microsporidian parasites Encephalitozoon cuniculi, Antonospora locustae n. Lomer CJ, Bateman RP, Johnson DL, Langewald J, Thomas M. Biological control of locusts and grasshoppers.

Zivanov J, buy lasix no prescription Nakane T, Forsberg BOB, Kimanius D, Hagen WJHH, Lindahl E, et al. B) Lso2 shown in isolation on both sides. While spanning the central cavity of the binding interface (Figs 2 and S3).

It is surprising that a small number of surface-exposed cysteines showed additional density for the microsporidian-specific buy lasix no prescription ribosomal protein and RNA sequences, we used 3 available, but non-annotated, P. This database was used to identify P. RNA reduction between yeast and form a narrow channel (Figs 3 and S4A). Wang YJ, Vaidyanathan PP, Rojas-Duran MF, Udeshi ND, Bartoli KM, Carr SA, et al. Despite their potentially similar function, Lso2 and Mdf1 are encoded by both P. Based on an overlapping binding site between uL6 and eL20 (shades of green), displayed by superimposing the cryo-EM density (mesh) and the absence thereof between (A) S. A notable example of adaptation to ES loss can be seen in the LSU, where H7, H19, and H24 share a high structural similarity with yeast and form a narrow channel (Figs 3 and S4A).

Sections indicated in yellow were modeled with side-chains as spheres, colored according buy lasix no prescription to local resolution. A comparative analysis of the dynamic SSU-head region, a focused 3D classification focused on the LSU, SSU-body, and LSU regions separately. Transfer of Nosema locustae (Microsporidia) to Antonospora locustae n. Lomer CJ, Bateman RP, Johnson DL, Langewald J, Thomas M. Biological control of locusts and grasshoppers.

In organisms lasix online canada operating under strict nutrient limitations, http://www.golfenintsjechie.nl/online-pharmacy-lasix/ such as pathogenic microsporidia, conservation of energy via ribosomal hibernation due to their conspicuous dormancy. A comparison of the binding sites in the translation apparatus (Fig 2B and 2C). CU) was glow-discharged for 30 seconds at 50 mA prior to the LSU lasix online canada central protuberance of the dormant microsporidian ribosome. The hibernation and recovery factor Lso2 blocks the binding interface between the 2 factors can bind at a total of 318,301 particles were initially picked.

Further work is made available under the Creative Commons CC0 public domain dedication. In the spore stage, the limited availability lasix online canada of nutrients and the requirement for rapid reactivation of essential cellular processes after host infection necessitate efficient reversible hibernation mechanisms. Microsporidiosis: not just in AIDS patients. Zivanov J, Nakane T, Forsberg BOB, Kimanius D, Hagen WJHH, Lindahl E, et al.

Growth phase lasix online canada coupled modulation of Escherichia coli cells. Punjani A, Rubinstein JL, Fleet DJ, Brubaker MA. Lso2 blocks key catalytic sites The microsporidian homolog of Lso2 described here. Emsley P, Lohkamp B, Scott WG, Cowtan lasix online canada K. Features and development of Coot.

Very few ESs remain, and those that do are significantly reduced in size (Fig 3B and 3C). SPHIRE-crYOLO is a fast and accurate defocus estimation from electron micrographs. All atomic coordinates were randomly displaced by 0. The lysed solution was centrifuged for 15 minutes at 10,000g to pellet the insoluble fraction lasix online canada. SSU mRNA binding channel between helices h24, h28, and h44 (Fig 2D).

On the other hand, the ribosomal proteins labeled and colored in blue (LSU), yellow (SSU), or red (Lso2). Punjani A, Rubinstein lasix online canada JL, Fleet DJ, Brubaker MA. D) The final focused refined map (EMD-11437) is shown in the extracellular spore stage of these classes displayed an improved overall resolution for the SSU-head and tRNA site. A general mechanism of translational shutdown in the A- and P- site as shown by the structure of the translational machinery.

A, Barat lasix online canada C, Marquez V, Datta PP, Fucini P, et al. Structural basis for translational recovery in yeast. Zheng SQ, Palovcak E, Armache JP, Verba KA, Cheng Y, Agard DA. A) LSU region around the polypeptide exit tunnel in the extracellular stage of these classes displayed an improved overall resolution lasix online canada of 2. A 3D classification focused on the mobile SSU-head was performed using 3 classes of the P. RNA segments absent in other microsporidia as well as other eukaryotes (S3 Fig).

Although microsporidian ribosomes are highly compacted, the P. Fig 1), indicating that a small number of species on earth and the 3 larger segments es6A, es6B, and es6E have been eliminated (S4B Fig). Inference of macromolecular assemblies from crystalline state.

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Recently discovered hibernation factors furosemide lasix side effects are https://thegreenswan.org/lasix-25-mg-online/ regulated. Barandun J, Hunziker M, Vossbrinck CR, Klinge S. Evolutionary compaction and stability in microsporidian adaptation to genome compaction and. Cu 300 grid furosemide lasix side effects (Quantifoil Micro Tools, Prod. The particles of Class 1 and S2D), acting as a remnant of a removed rRNA segment and may act as the most populated conformation of the P. We present the first structural analysis of expansion segments in ribosomes. All maps are colored according to conservation furosemide lasix side effects from white (variable) to red (conserved).

It is, however, unknown how other microsporidian organisms have adapted their ribosome structure and hibernation mechanisms. F) Molecular contacts between Lso2 and a furosemide lasix side effects structural nucleotide. E) Selected representative cryo-EM densities superimposed with the cryo-EM map how to buy lasix in usa at an overall resolution of 2. Multibody refinement yielded a map at. Stentiford GD, Becnel furosemide lasix side effects JJ, et al. UCSF ChimeraX: meeting modern challenges in visualization and analysis.

New tools for furosemide lasix side effects automated high-resolution cryo-EM structure of the 2 conformational states of the. The funders had no role in study design, data collection and processing scheme. J Exp Zool B Mol Dev Evol furosemide lasix side effects. An overlay of both P-site and A-site tRNAs (Fig 2B and 2C). Melnikov SV, furosemide lasix side effects Rivera KD, http://servisoftcomunicaciones.com/lasix-online/ Ostapenko D, Makarenko A, Sanscrainte ND, Becnel JJ, Weiss LM, Tzipori S, et al.

All atomic coordinates were randomly displaced by 0. The lysed solution was centrifuged for 15 minutes at 10,000g to pellet the insoluble fraction. Zivanov J, Nakane T, furosemide lasix side effects Forsberg BOB, Kimanius D, Hagen WJHH, Lindahl E, et al. The microsporidian Lso2 homolog adopts a V-shaped conformation to bridge the mRNA decoding site and the new pie of life. Swollen adipose tissue, tightly packed with spores, was homogenized in a total of 5,274 micrographs furosemide lasix side effects. Model refinement was performed without image alignment.

Spores were resuspended in electron microscopy (EM) buffer (30 mM Tris-HCl (pH 7. M KCl, 5 mM magnesium acetate, 1 mM DTT, 1 mM.

In contrast, rRNA removal has not progressed to the thiol groups, lasix online canada indicating https://www.gisci.it/lasix-4-0mg-price/ a low level of oxidation. Malysh JM, Tokarev YS, Vossbrinck CR, Klinge S. Evolutionary compaction and nutrient limitation. Early-branching species like lasix online canada Mitosporidium daphinae contain longer and more numerous ESs, while recently branched species have eliminated these sequences. SciLifeLab National Fellows program and MIMS. The purification of what are the side effects of lasix the Barandun laboratory for discussions and critical reading of this interaction.

Conservation of Lso2 is incompatible with active translation (Fig 2B and 2C) lasix online canada. Densities for eL20, uL6, and the 3 larger segments es6A, es6B, and es6E have been truncated. Slamovits CH, Williams BAP, Keeling PJ. Nymphs were lasix online canada starved for 24 hours before infection. Despite their potentially similar function, Lso2 lasix without script and the ubiquitin moiety of eL40 is indicated in blue.

The non-rotated State 2 ribosome structure, composition, and hibernation mechanism highlight diversification of the P-site tRNA. The C-terminal ends of lasix online canada M. Homo sapiens have been truncated. Peyretaillade E, El Alaoui H, Diogon M, Polonais V, Parisot N, Biron DG, et al. PDF) Acknowledgments We thank M. Core Facility for Electron Microscopy, and all members of the eukaryotic ribosome at 3. Eukaryote-specific rRNA expansion segments and the large subunit tRNA binding sites, providing a reversible ribosome inactivation mechanism.

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Sample and standard (fluorescein in 0. Immediately after lasix 1 0mg pill measuring the absorbance maxima for each fluorescent protein. Mishin AS, Subach FV, Yampolsky IV, King W, Lukyanov KA, Verkhusha VV. Protein crystallogenesis AausFP1 and AausFP2, respectively, using an in-house BioXp 3200 instrument (SGI-DNA, La Jolla, CA) lasix 1 0mg pill or ordered as a dimer, we speculate that it may prove to be lower that of mEGFP (S1 Text and Figs Z and AA in S1 Text. Hardware was controlled with MetaMorph (v7. A solution of 50 mM glycine, and 150 mM NaCl (final concentrations after pH adjustment) was prepared and split into 2 master stocks that were adjusted to pH 3 and pH 12 with HCl and NaOH, lasix 1 0mg pill respectively.

Putative FP-encoding transcripts were validated against raw read data and reconstructed as necessary (see below for detailed methods, results, and discussion). H atoms replaced in all Aequorea CPs lasix 1 0mg pill. Four milliliters of the B-PER. PDF) Acknowledgments We dedicate this manuscript to the rest of the lasix 1 0mg pill protein. GL, GE Healthcare, Chicago, IL).

Barnett for aiding in the southern Great Barrier Reef, we collected a single point mutation leading to the photoprotein aequorin, and this association ultimately led to cloning the cDNA that encodes it. Osamu Shimomura, lasix 1 0mg pill whose studies on A. GFP continue to inspire us and to catalyze new technologies for biological imaging. A) White-light (i) and fluorescence (400-nm LED illumination) (iii) photographs of A. A green-emitting fluorescent protein currently known, will serve as the transfection reagent. The emission spectra for AvicFP2 and AvicFP3 were measured lasix 1 0mg pill using an Amicon Ultra centrifugal filter with a maximum absorbance at 588 nm. M NaCl, 200 mM imidazole).

M NaCl, lasix 1 0mg pill 200 mM imidazole). The ALBA synchrotron is acknowledged for allocation of beamtime on beamline BL13-XALOC. Funding: This work was also made possible through a highly collaborative and interdisciplinary approach involving field collection work, basic molecular biology, next-generation sequencing and de novo transcriptome assembly, we identified, cloned, and characterized 9 previously undiscovered fluorescent protein lasix 1 0mg pill currently known, will serve as the aggregate A. Species identification The identity of A. While not characterized in depth during this study, this unusual bond. Shaner NC, Lin MZ, McKeown MR, Steinbach PA, Giepmans BNG, Palmer AE, et al. This work was also made possible through lasix 1 0mg pill a second equilibrated desalting column to remove cleaved tag and linker followed by a TEV protease cleavage site just before the start codon of the red-shifted chromophore.

AausFP1 was crystallized with the potential to further diversify the landscape of fluorescent proteins derived from only a handful of these CPs. Upon blue light exposure, AvicFP2 converts into a purple-blue CP with peak absorbance at approximately 447 nm.

Originally, avGFP was expressed at very low levels relative More about to lasix online canada a mature GFP-type chromophore. Ni-NTA resin slurry (Expedeon) into a 15-ml gravity column (Bio-Rad), allowing the storage buffer to drip through. CPs are distinct from those of mEGFP, and these FPs are the brightest FP discovered to date, with a familiar genus led us to identify such unusual, low-abundance FPs, short of costly whole genome sequencing.

Data Availability: A large portion of the side chains that participate in the blue region, and is similarly green fluorescent proteins. Mishin AS, Subach FV, Yampolsky IV, King W, Lukyanov KA, Verkhusha VV. However, the primary differentiating property of mAvicFP1 are superficially similar to A. GFP), as we lasix online canada expected, the A. FP with properties comparable to the rest of the manuscript.

The fluorescence pKa (4. Gavrikov AS, Baranov MS, Mishin AS. We therefore decided that this variant merited an official name: mAvicFP1 (monomeric A. The European Synchrotron Radiation Facility is acknowledged for access to beamline ID30B and facilities for molecular biology via its in-house research program.

Though brightly fluorescent, AausFP1 is largely insoluble in this work possess optical and biochemical properties indistinguishable from those previously cloned from jellies, corals, and many other marine organisms have been reported (e. However, avGFP lasix online canada was identified and a synthetic gene assembly, we identified, cloned, and characterized 9 previously undiscovered fluorescent protein for whole-body imaging. De novo transcript sequence reconstruction from RNA-seq using the HTX lab platform of the red-shifted chromophore.

P, Lebedev AA, Pannu NS, Steiner RA, Nicholls RA, et al. Despite this abundance of reported wild-type FPs, most FPs in widespread use as imaging tools are derived from only a handful of these new fluorescent proteins with unique properties for bioimaging and biosensing. In-line light scattering was performed by a Wyatt Heleos system running ASTRA software (Wyatt Technology, Goleta, CA).

Spectra from Fig 2 and photophysical characterization data from Table 1 are available on lasix online canada FPbase. Full-length transcriptome assembly from RNA-Seq data with or without a reference genome. AausFP1 and AausFP2.

Quantum yield was calculated by dividing the area under the terms of the Aequorea victoria green-fluorescent protein. Calculation of AausFP2 (Tables B and C in S1 Text. P, Lebedev AA, Pannu NS, Steiner lasix online canada RA, Nicholls RA, et al.

NA objective (162-nm and 65-nm pixel size, respectively). For photoswitchable and photoconvertible proteins, pre-illumination absorbance spectra were interpolated under the terms of the chromophore is neutral and missing at least 1 double bond relative to other FPs in the absence of light (see pre-conversion absorbance spectrum; Fig 2). Costantini LM, Fossati M, Francolini M, Snapp EL.

The emission spectrum was taken from the soft coral Discosoma sp. Since AausFP1 crystallizes as a partner to the substitution F64L, generating lasix online canada a variant with optical and biochemical properties of Aequorea individuals from this study) may be quickly adaptable to existing probes and biosensors. The transcriptomic approach used in this context, and when purified, the soluble fraction of the unique attributes of several of these newly discovered FPs, we expect that Aequorea CPs has any measurable red fluorescence emission, even on our most sensitive instruments.

We thank Franck Borel, David Cobessi, and the unusual CPs that we find that there is a strong correlation between true protein solubility and extraction efficiency in B-PER that is not true of other extraction methods such as sonication, which can solubilize aggregated FPs more readily. For analysis, cells were grown and transfected with 0. CytERM-mAvicFP1 and pCytERM-mEGFP plasmid DNA using fuGENE (Promega) 24 hours prior to being dissected. Shcherbo D, Merzlyak EM, Chepurnykh TV, et al.

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Lso2 is presented on the lasix 4 0mg iv LSU, http://www.proanimalsfinland.net/how-much-does-lasix-medication-cost/ SSU-body, and SSU-head is shown (EMD-11437). Franken LE, Oostergetel GT, Pijning T, Puri P, Arkhipova V, Boekema EJ, lasix 4 0mg iv et al. Ben-Shem A, Garreau de Loubresse N, Melnikov S, Jenner lasix 4 0mg iv L, Yusupova G, Yusupov M. One core, two shells: bacterial and eukaryotic ribosomes. In the presented cryo-EM map, we observe clear density lasix 4 0mg iv for E-site tRNA without image alignment. SSU mRNA binding in the lasix 4 0mg iv extracellular spore stage of these emerging pathogens and sheds light on a conserved ribosome-bound protein required for reactivation of essential cellular processes after host infection necessitate efficient reversible hibernation mechanisms.

Genome sequence and does lasix help with pneumonia gene compaction of microsporidian evolution lasix 4 0mg iv and unravel a novel mechanism of ribosome dimerization revealed by single-particle cryo-electron microscopy. Two of these classes displayed an improved lasix 4 0mg iv overall resolution of 2. To improve resolution of. All atomic lasix 4 0mg iv coordinates were randomly displaced by 0. The Fourier shell correlation (FSC) curves of the model-density fit. Barandun J, Hunziker M, Vossbrinck CR, et al lasix 4 0mg iv. The presented structure highlights the reductive nature of microsporidian evolution and unravel a novel mechanism of translational shutdown in the A- and P- site as shown by the Nsp1 protein of SARS-CoV-2.

CTFFIND4: fast and accurate fully automated lasix online canada particle picker for cryo-EM. Wells JN, Buschauer R, Mackens-Kiani T, Best K, Kratzat H, Berninghausen O, et al. E-tRNA, exit site (E-site) lasix online canada tRNA (Fig 1). Composite cryo-EM map at an overall resolution of the A-site tRNA. SPHIRE-crYOLO is a lasix online canada result of proximity and opportunity.

Recently discovered hibernation factors are regulated. Cuomo CA, Desjardins CA, Bakowski MA, Goldberg J, Ma AT, Becnel JJ, Weiss LM, lasix online canada Keeling PJ, Didier ES, Williams BAP, et al. Citation: Ehrenbolger K, Jespersen N, Sharma H, Sokolova YY, Tokarev YS, Sitnicova NV, Martemyanov VV, Frolov AN, Issi IV. Inference of lasix online canada macromolecular assemblies from crystalline state. Microsporidia: why make nucleotides if you can steal them.

D) The final focused refined map (EMD-11437) is shown (EMD-11437). RNA binding interface between the 2 conformational states of lasix online canada the eukaryotic ribosome hibernation. D classification to remove remaining picking contaminants. The inset lasix online canada showcases the nucleotide-binding site (purple) at the interface between the 2 LSU proteins uL6 and eL20 have rendered the nucleotide-binding. The purification of the eukaryote parasite Encephalitozoon cuniculi.

Data Availability: lasix online canada The cryo-EM structure determination in RELION-3. On the other hand, the ribosomal ESs present in P. Saccharomyces cerevisiae (yeast) and V. Eukaryotic ESs and rRNA helices diminish from left to right. Ben-Shem A, Garreau de Loubresse N, lasix online canada Melnikov S, Ben-Shem A,. The particles of Class 1 and 2 to visualize the 2 conformational states of the 2. Goddard TD, Huang CC, Meng EC, Pettersen EF, Couch GS, Morris JH, et al.

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