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- Cossarizza, A., et al.
(författare)
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Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition)
- 2019
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Ingår i: European Journal of Immunology. - : Wiley. - 0014-2980 .- 1521-4141. ; 49:10, s. 1457-1973
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Tidskriftsartikel (refereegranskat)abstract
- These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer-reviewed by leading experts in the field, making this an essential research companion.
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2. |
- Singh, B. P., et al.
(författare)
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Experimental access to Transition Distribution Amplitudes with the PANDA experiment at FAIR
- 2015
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Ingår i: European Physical Journal A. Hadrons and Nuclei. - : Springer Science and Business Media LLC. - 1434-6001 .- 1434-601X. ; 51:8
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Tidskriftsartikel (refereegranskat)abstract
- Baryon-to-meson Transition Distribution Amplitudes (TDAs) encoding valuable new information on hadron structure appear as building blocks in the collinear factorized description for several types of hard exclusive reactions. In this paper, we address the possibility of accessing nucleon-to-pion (pi N) TDAs from (p) over barp -> e(+)e(-)pi(0) reaction with the future PANDA detector at the FAIR facility. At high center-of-mass energy and high invariant mass squared of the lepton pair q(2), the amplitude of the signal channel (p) over barp -> e(+)e(-)pi(0) admits a QCD factorized description in terms of pi N TDAs and nucleon Distribution Amplitudes (DAs) in the forward aid backward kinematic regimes. Assuming the validity of this factorized description, we perform feasibility studies for measuring (p) over barp -> e(+)e(-)pi(0) with the PANDA detector. Detailed simulations on signal reconstruction efficiency as well as on rejection of the most severe background channel, i.e. (p) over barp -> pi(+)pi(-)pi(0) were performed for the center-of-mass energy squared s = 5 GeV2 and s = 10 GeV2, in the kinematic regions 3.0 < q(2) < 4.3 GeV2 and 5 < q(2) < 9 GeV2, respectively, with a neutral pion scattered in the forward or backward cone vertical bar cos theta(pi 0)vertical bar > 0.5 in the proton-antiproton center-of-mass frame. Results of the simulation show that the particle identification capabilities of the PANDA detector will allow to achieve a background rejection factor of 5 . 10(7) (1 . 10(7)) at low (high) q(2) for s = 5 GeV2, and of 1 . 10(8) (6 . 10(6)) at low (high) q(2) for s = 10 GeV2, while keeping the signal reconstruction efficiency at around 40%. At both energies, a clean lepton signal can be reconstructed with the expected statistics corresponding to 2 of integrated luminosity. The cross sections obtained from the simulations are used to show that a test of QCD collinear factorization can be done at the lowest order by measuring scaling laws and angular distributions. The future measurement of the signal channel cross section with PANDA will provide a new test of the perturbative QCD description of a novel class of hard exclusive reactions and will open the possibility of experimentally accessing pi N TDAs.
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3. |
- Wendisch, M., et al.
(författare)
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Atmospheric and Surface Processes, and Feedback Mechanisms Determining Arctic Amplification: A Review of First Results and Prospects of the (AC)(3) Project
- 2023
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Ingår i: Bulletin of the American Meteorological Society. - : American Meteorological Society. - 0003-0007 .- 1520-0477. ; 104:1
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Tidskriftsartikel (refereegranskat)abstract
- Mechanisms behind the phenomenon of Arctic amplification are widely discussed. To contribute to this debate, the (AC)(3) project was established in 2016 (www.ac3-tr.de/). It comprises modeling and data analysis efforts as well as observational elements. The project has assembled a wealth of ground-based, airborne, shipborne, and satellite data of physical, chemical, and meteorological properties of the Arctic atmosphere, cryosphere, and upper ocean that are available for the Arctic climate research community. Short-term changes and indications of long-term trends in Arctic climate parameters have been detected using existing and new data. For example, a distinct atmospheric moistening, an increase of regional storm activities, an amplified winter warming in the Svalbard and North Pole regions, and a decrease of sea ice thickness in the Fram Strait and of snow depth on sea ice have been identified. A positive trend of tropospheric bromine monoxide (BrO) column densities during polar spring was verified. Local marine/biogenic sources for cloud condensation nuclei and ice nucleating particles were found. Atmospheric-ocean and radiative transfer models were advanced by applying new parameterizations of surface albedo, cloud droplet activation, convective plumes and related processes over leads, and turbulent transfer coefficients for stable surface layers. Four modes of the surface radiative energy budget were explored and reproduced by simulations. To advance the future synthesis of the results, cross-cutting activities are being developed aiming to answer key questions in four focus areas: lapse rate feedback, surface processes, Arctic mixed-phase clouds, and airmass transport and transformation.
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6. |
- Olahova, M., et al.
(författare)
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POLRMT mutations impair mitochondrial transcription causing neurological disease
- 2021
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- While >300 disease-causing variants have been identified in the mitochondrial DNA (mtDNA) polymerase gamma, no mitochondrial phenotypes have been associated with POLRMT, the RNA polymerase responsible for transcription of the mitochondrial genome. Here, we characterise the clinical and molecular nature of POLRMT variants in eight individuals from seven unrelated families. Patients present with global developmental delay, hypotonia, short stature, and speech/intellectual disability in childhood; one subject displayed an indolent progressive external ophthalmoplegia phenotype. Massive parallel sequencing of all subjects identifies recessive and dominant variants in the POLRMT gene. Patient fibroblasts have a defect in mitochondrial mRNA synthesis, but no mtDNA deletions or copy number abnormalities. The in vitro characterisation of the recombinant POLRMT mutants reveals variable, but deleterious effects on mitochondrial transcription. Together, our in vivo and in vitro functional studies of POLRMT variants establish defective mitochondrial transcription as an important disease mechanism. POLRMT is key for transcription of the mitochondrial genome, yet has not been implicated in mitochondrial disease to date. Here, the authors identify mutations in POLRMT in individuals with mitochondrial disease-related phenotypes and characterise underlying defects in mitochondrial transcription.
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9. |
- Sparks, Justin L, et al.
(författare)
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RNase H2-Initiated Ribonucleotide Excision Repair
- 2012
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 47:6, s. 980-986
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Tidskriftsartikel (refereegranskat)abstract
- Ribonucleotides are incorporated into DNA by the replicative DNA polymerases at frequencies of about 2 per kb, which makes them by far the most abundant form of potential DNA damage in the cell. Their removal is essential for restoring a stable intact chromosome. Here, we present a complete biochemical reconstitution of the ribonucleotide excision repair (RER) pathway with enzymes purified from Saccharomyces cerevisiae. RER is most efficient when the ribonucleotide is incised by RNase H2, and further excised by the flap endonuclease FEN1 with strand displacement synthesis carried out by DNA polymerase δ, the PCNA clamp, its loader RFC, and completed by DNA ligase I. We observed partial redundancy for several of the enzymes in this pathway. Exo1 substitutes for FEN1 and Pol ε for Pol δ with reasonable efficiency. However, RNase H1 fails to substitute for RNase H2 in the incision step of RER.
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10. |
- Vieira, Natassia M., et al.
(författare)
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Jagged 1 Rescues the Duchenne Muscular Dystrophy Phenotype
- 2015
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Ingår i: Cell. - : Elsevier BV. - 0092-8674 .- 1097-4172. ; 163:5, s. 1204-1213
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Tidskriftsartikel (refereegranskat)abstract
- Duchenne muscular dystrophy (DMD), caused by mutations at the dystrophin gene, is the most common form of muscular dystrophy. There is no cure for DMD and current therapeutic approaches to restore dystrophin expression are only partially effective. The absence of dystrophin in muscle results in dysregulation of signaling pathways, which could be targets for disease therapy and drug discovery. Previously, we identified two exceptional Golden Retriever muscular dystrophy (GRMD) dogs that are mildly affected, have functional muscle, and normal lifespan despite the complete absence of dystrophin. Now, our data on linkage, whole-genome sequencing, and transcriptome analyses of these dogs compared to severely affected GRMD and control animals reveals that increased expression of Jagged1 gene, a known regulator of the Notch signaling pathway, is a hallmark of the mild phenotype. Functional analyses demonstrate that Jagged1 overexpression ameliorates the dystrophic phenotype, suggesting that Jagged1 may represent a target for DMD therapy in a dystrophin-independent manner.
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11. |
- Nick McElhinny, Stephanie A, et al.
(författare)
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Abundant ribonucleotide incorporation into DNA by yeast replicative polymerases.
- 2010
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 107:11, s. 4949-4954
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Tidskriftsartikel (refereegranskat)abstract
- Measurements of nucleoside triphosphate levels in Saccharomyces cerevisiae reveal that the four rNTPs are in 36- to 190-fold molar excess over their corresponding dNTPs. During DNA synthesis in vitro using the physiological nucleoside triphosphate concentrations, yeast DNA polymerase epsilon, which is implicated in leading strand replication, incorporates one rNMP for every 1,250 dNMPs. Pol delta and Pol alpha, which conduct lagging strand replication, incorporate one rNMP for every 5,000 or 625 dNMPs, respectively. Discrimination against rNMP incorporation varies widely, in some cases by more than 100-fold, depending on the identity of the base and the template sequence context in which it is located. Given estimates of the amount of replication catalyzed by Pols alpha, delta, and epsilon, the results are consistent with the possibility that more than 10,000 rNMPs may be incorporated into the nuclear genome during each round of replication in yeast. Thus, rNMPs may be the most common noncanonical nucleotides introduced into the eukaryotic genome. Potential beneficial and negative consequences of abundant ribonucleotide incorporation into DNA are discussed, including the possibility that unrepaired rNMPs in DNA could be problematic because yeast DNA polymerase epsilon has difficulty bypassing a single rNMP present within a DNA template.
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- Schymanski, Emma L, et al.
(författare)
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Non-target screening with high-resolution mass spectrometry : critical review using a collaborative trial on water analysis
- 2015
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Berlin/Heidelberg. - 1618-2642 .- 1618-2650. ; 407:21, s. 6237-6255
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Forskningsöversikt (refereegranskat)abstract
- In this article, a dataset from a collaborative non-target screening trial organised by the NORMAN Association is used to review the state-of-the-art and discuss future perspectives of non-target screening using high-resolution mass spectrometry in water analysis. A total of 18 institutes from 12 European countries analysed an extract of the same water sample collected from the River Danube with either one or both of liquid and gas chromatography coupled with mass spectrometry detection. This article focuses mainly on the use of high resolution screening techniques with target, suspect, and non-target workflows to identify substances in environmental samples. Specific examples are given to emphasise major challenges including isobaric and co-eluting substances, dependence on target and suspect lists, formula assignment, the use of retention information, and the confidence of identification. Approaches and methods applicable to unit resolution data are also discussed. Although most substances were identified using high resolution data with target and suspect-screening approaches, some participants proposed tentative non-target identifications. This comprehensive dataset revealed that non-target analytical techniques are already substantially harmonised between the participants, but the data processing remains time-consuming. Although the objective of a "fully-automated identification workflow" remains elusive in the short term, important steps in this direction have been taken, exemplified by the growing popularity of suspect screening approaches. Major recommendations to improve non-target screening include better integration and connection of desired features into software packages, the exchange of target and suspect lists, and the contribution of more spectra from standard substances into (openly accessible) databases.
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13. |
- Watt, Danielle L, et al.
(författare)
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Replication of ribonucleotide-containing DNA templates by yeast replicative polymerases
- 2011
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Ingår i: DNA Repair. - : Elsevier BV. - 1568-7864 .- 1568-7856. ; 10:8, s. 897-902
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Tidskriftsartikel (refereegranskat)abstract
- The major replicative DNA polymerases of S. cerevisiae (Pols α, δ, and ɛ) incorporate substantial numbers of ribonucleotides into DNA during DNA synthesis. When these ribonucleotides are not removed in vivo, they reside in the template strand used for the next round of replication and could potentially reduce replication efficiency and fidelity. To examine if the presence of ribonucleotides in a DNA template impede DNA synthesis, we determined the efficiency with which Pols α, δ, and ɛ copy DNA templates containing a single ribonucleotide. All three polymerases can replicate past ribonucleotides. Relative to all-DNA templates, bypass of ribo-containing templates is slightly reduced, to extents that depend on the identity of the ribo and the sequence context in which it resides. Bypass efficiencies for Pols δ and ɛ were increased by increasing the dNTP concentrations to those induced by cellular stress, and in the case of Pol ɛ, by inactivating the 3'-exonuclease activity. Overall, ribonucleotide bypass efficiencies are comparable to, and usually exceed, those for the common oxidative stress-induced lesion 8-oxo-guanine.
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