scholarly journals Towards high spatially resolved microproteomics using expansion microscopy

2021 ◽  
Author(s):  
Lauranne Drelich ◽  
Soulaimane Aboulouard ◽  
Julien Franck ◽  
Michel Salzet ◽  
Isabelle Fournier ◽  
...  
Keyword(s):  
Tissue Section ◽  
Large Scale ◽  
Protein Identification ◽  
Tissue Expansion ◽  
Proteinase K ◽  
Protein Diffusion ◽  
Morphological Examination ◽  
Spatially Resolved ◽  
Cell Structures ◽  
Proteomic Study

AbstractExpansion microscopy is an emerging approach for morphological examination of biological specimens at nanoscale resolution using conventional optical microscopy. To achieve physical separation of cell structures, tissues are embedded in a swellable polymer and expanded several folds in an isotropic manner. This work shows the development and optimization of physical tissue expansion as a new method for spatially resolved large scale proteomics. Herein, we established a novel method to enlarge the tissue section to be compatible with manual microdissection on regions of interest and to carry out MS-based proteomic analysis. A major issue in the Expansion microscopy is the loss of proteins information during the mechanical homogenization phase due to the use of Proteinase K. For isotropic expansion, different homogenization agents are investigated, both to maximize protein identification and to minimize protein diffusion. Better results are obtained with SDS. From a tissue section enlarge more than 3-fold, we have been able to manually cut out regions of 1mm in size, equivalent to 300µm in their real size. We identified up to 655 proteins from a region corresponding to an average of 940 cells. This approach can be performed easily without any expensive sampling instrument. We demonstrated the compatibility of sample preparation for expansion microscopy and proteomic study in a spatial context.Abstract graphic

scholarly journals Rapid, large-scale species discovery in hyperdiverse taxa using 1D MinION sequencing

BMC Biology ◽  
2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Amrita Srivathsan ◽  
Emily Hartop ◽  
Jayanthi Puniamoorthy ◽  
Wan Ting Lee ◽  
Sujatha Narayanan Kutty ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Large Scale ◽  
Low Cost ◽  
Small Body ◽  
Small Subset ◽  
Similar Species ◽  
Large Species ◽  
Morphological Examination ◽  
Species Discovery ◽  
Short Period

Abstract Background More than 80% of all animal species remain unknown to science. Most of these species live in the tropics and belong to animal taxa that combine small body size with high specimen abundance and large species richness. For such clades, using morphology for species discovery is slow because large numbers of specimens must be sorted based on detailed microscopic investigations. Fortunately, species discovery could be greatly accelerated if DNA sequences could be used for sorting specimens to species. Morphological verification of such “molecular operational taxonomic units” (mOTUs) could then be based on dissection of a small subset of specimens. However, this approach requires cost-effective and low-tech DNA barcoding techniques because well-equipped, well-funded molecular laboratories are not readily available in many biodiverse countries. Results We here document how MinION sequencing can be used for large-scale species discovery in a specimen- and species-rich taxon like the hyperdiverse fly family Phoridae (Diptera). We sequenced 7059 specimens collected in a single Malaise trap in Kibale National Park, Uganda, over the short period of 8 weeks. We discovered > 650 species which exceeds the number of phorid species currently described for the entire Afrotropical region. The barcodes were obtained using an improved low-cost MinION pipeline that increased the barcoding capacity sevenfold from 500 to 3500 barcodes per flowcell. This was achieved by adopting 1D sequencing, resequencing weak amplicons on a used flowcell, and improving demultiplexing. Comparison with Illumina data revealed that the MinION barcodes were very accurate (99.99% accuracy, 0.46% Ns) and thus yielded very similar species units (match ratio 0.991). Morphological examination of 100 mOTUs also confirmed good congruence with morphology (93% of mOTUs; > 99% of specimens) and revealed that 90% of the putative species belong to the neglected, megadiverse genus Megaselia. We demonstrate for one Megaselia species how the molecular data can guide the description of a new species (Megaselia sepsioides sp. nov.). Conclusions We document that one field site in Africa can be home to an estimated 1000 species of phorids and speculate that the Afrotropical diversity could exceed 200,000 species. We furthermore conclude that low-cost MinION sequencers are very suitable for reliable, rapid, and large-scale species discovery in hyperdiverse taxa. MinION sequencing could quickly reveal the extent of the unknown diversity and is especially suitable for biodiverse countries with limited access to capital-intensive sequencing facilities.

Genetic and morphological diversity of the cosmopolitan chaetognath Pseudosagitta maxima (Conant, 1896) in the Atlantic Ocean and its relationship with the congeneric species

2017 ◽  
Vol 74 (7) ◽  
pp. 1875-1884 ◽  
Author(s):  
Dmitry N. Kulagin ◽  
Tatiana V. Neretina
Keyword(s):  
Atlantic Ocean ◽  
Large Scale ◽  
Phylogenetic Analyses ◽  
Morphological Diversity ◽  
Maturity Stage ◽  
Congeneric Species ◽  
Morphological Examination ◽  
Posterior Teeth ◽  
Biogeographic Patterns ◽  
True Diversity

Abstract Until recently many oceanic zooplankton species have been considered as cosmopolitan organisms. At present it became evident that some of them comprise many distinct molecular operational taxonomic units (MOTUs) that often are regarded as cryptic species. As they can significantly change our perceptions of large-scale biogeographic patterns, it is important to characterize the true diversity within common and ecologically important groups. We have analysed the molecular and morphological diversity of the cosmopolitan mesopelagic chaetognath Pseudosagitta maxima throughout the Atlantic Ocean from 60° S to 85° N and its position within the genus Pseudosagitta. Three distinct mitochondrial clades within P. maxima were revealed with phylogenetic analyses (Maximum Likelihood, Bayesian Inference) and were geographically separated. The subsequent analyses of nuclear markers (H3, ITS1) have shown that P. maxima most likely comprises two distinct MOTUs, tropical and bipolar, that also have some morphological differences. The latter MOTU consists of two genetically slightly divergent populations: southern and northern. The morphological examination allowed the determination of a character (type of hook coloration) that accurately distinguishes juveniles of the P. maxima complex from the other congeneric species. Molecular data have shown that evolutionary P. lyra and P. gazellae are more closely related to each other than to P. maxima. Number of hooks, number of anterior and posterior teeth and the arrangement of ova in the ovary were proposed to be the most useful morphological characters to distinguish between tropical and bipolar MOTUs within the P. maxima complex. The first three characters should be determined for each maturity stage separately.

scholarly journals Survey of Cattle in Northeast Colorado for Evidence of Chronic Wasting Disease: Geographical and High-Risk Targeted Sample

2003 ◽  
Vol 15 (3) ◽  
pp. 274-277 ◽  
Author(s):  
Daniel H. Gould ◽  
James L. Voss ◽  
Michael W. Miller ◽  
Annette M. Bachand ◽  
Bruce A. Cummings ◽  
...  
Keyword(s):  
High Risk ◽  
Large Scale ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Proteinase K ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Adult Cattle ◽  
Endemic Areas ◽  
Geographically Targeted

A geographically targeted survey of potentially high-risk, adult cattle in chronic wasting disease (CWD)–endemic areas in Colorado was initiated to assess the possibility of the spread of CWD from deer to cattle under natural conditions. Surveyed cattle were sympatric with free-roaming deer in geographically defined areas where CWD occurs and where CWD prevalence has been estimated. To qualify for inclusion in the survey, cattle had to be at least 4 years old and had to have spent a minimum of 4 years in surveyed areas. Brains from culled cattle were examined microscopically and immunohistochemically for tissue alterations indicative of a transmissible spongiform encephalopathy (TSE). Two hundred sixty-two brains were suitable for evaluation and were found to lack changes indicative of a TSE infection. Prion deposition was not demonstrable using a method involving formic acid and proteinase-K treatment before application of monoclonal antibody to bovine prion protein (F99/97.6.1). Some incidental neuropathologic changes unrelated to those of TSEs were detected. Findings from this study suggest that large-scale spread of CWD from deer to cattle under natural range conditions in CWD-endemic areas of northeast Colorado is unlikely.

Sex differences in white adipose tissue expansion: emerging molecular mechanisms

2021 ◽  
Vol 135 (24) ◽  
pp. 2691-2708
Author(s):  
Simon T. Bond ◽  
Anna C. Calkin ◽  
Brian G. Drew
Keyword(s):  
Sex Differences ◽  
Sexual Dimorphism ◽  
Gene Networks ◽  
Large Scale ◽  
Molecular Mechanisms ◽  
Association Studies ◽  
Tissue Expansion ◽  
Economic Systems ◽  
Genomic Association ◽  
Males And Females

Abstract The escalating prevalence of individuals becoming overweight and obese is a rapidly rising global health problem, placing an enormous burden on health and economic systems worldwide. Whilst obesity has well described lifestyle drivers, there is also a significant and poorly understood component that is regulated by genetics. Furthermore, there is clear evidence for sexual dimorphism in obesity, where overall risk, degree, subtype and potential complications arising from obesity all differ between males and females. The molecular mechanisms that dictate these sex differences remain mostly uncharacterised. Many studies have demonstrated that this dimorphism is unable to be solely explained by changes in hormones and their nuclear receptors alone, and instead manifests from coordinated and highly regulated gene networks, both during development and throughout life. As we acquire more knowledge in this area from approaches such as large-scale genomic association studies, the more we appreciate the true complexity and heterogeneity of obesity. Nevertheless, over the past two decades, researchers have made enormous progress in this field, and some consistent and robust mechanisms continue to be established. In this review, we will discuss some of the proposed mechanisms underlying sexual dimorphism in obesity, and discuss some of the key regulators that influence this phenomenon.

scholarly journals KK-DBP: A Multi-Feature Fusion Method for DNA-Binding Protein Identification Based on Random Forest

2021 ◽  
Vol 12 ◽  
Author(s):  
Yuran Jia ◽  
Shan Huang ◽  
Tianjiao Zhang
Keyword(s):  
Dna Binding ◽  
Prediction Accuracy ◽  
Large Scale ◽  
Protein Identification ◽  
Feature Fusion ◽  
Binding Protein ◽  
Rapid Development ◽  
Dna Binding Protein ◽  
Feature Extraction Method ◽  
Independent Test Dataset

DNA-binding protein (DBP) is a protein with a special DNA binding domain that is associated with many important molecular biological mechanisms. Rapid development of computational methods has made it possible to predict DBP on a large scale; however, existing methods do not fully integrate DBP-related features, resulting in rough prediction results. In this article, we develop a DNA-binding protein identification method called KK-DBP. To improve prediction accuracy, we propose a feature extraction method that fuses multiple PSSM features. The experimental results show a prediction accuracy on the independent test dataset PDB186 of 81.22%, which is the highest of all existing methods.

scholarly journals Recent developments and prospects in the composition, extraction, stability, delivery system, digestion and food applications of plant oil bodies

2021 ◽  
Author(s):  
Jia Hao ◽  
Duoxia Xu ◽  
Yangping Cao
Keyword(s):  
Delivery System ◽  
Large Scale ◽  
Protein Identification ◽  
Meat Products ◽  
Calorie Intake ◽  
Oil Bodies ◽  
Plant Oil ◽  
Electrostatic Deposition ◽  
Food Applications ◽  
The Stability

Oil bodies (OBs) are micron- or submicron-sized sub-organelles widely found in plants seeds and nuts. The structure OBs is composed of a core of triglycerides covered by a phospholipid-protein layer, which ensures the stability of the OBs under extreme environmental conditions and further protects core lipids as energy reserves. As naturally pre-emulsified oil-in-water emulsions, OBs have been gradually applied to replace synthetically engineered oil droplets. In this paper, the recent research on the composition, extraction, stability, delivery system, digestion, food applications and future perspectives of plant OBs are reviewed. Recent studies have focused on the OBs surface protein identification and function, large-scale extraction techniques such as enzyme assisted, high pressure, ultrasound, and extrusion and the reconstituted OBs. Electrostatic deposition of polysaccharides significantly improves the stability of OBs emulsions. OBs emulsions have promising applications to encapsulate bioactive compounds, deliver targeted drugs, and prepare gels and edible functional films. The digestive behavior of OBs emulsions is similar to that of protein-stabilized emulsions, which can increase the satiety, effectively help reduce calorie intake and improve the bioavailability of functional factors. It has also promoted the development of simulated dairy, spices and meat products.

scholarly journals 3D mapping of host-parasite-microbiome interactions reveals metabolic determinants of tissue tropism and disease tolerance in Chagas disease

2019 ◽  
Author(s):  
Ekram Hossain ◽  
Sharmily Khanam ◽  
Chaoyi Wu ◽  
Sharon Lostracco-Johnson ◽  
Diane Thomas ◽  
...  
Keyword(s):  
Chagas Disease ◽  
Large Intestine ◽  
Large Scale ◽  
Scale Effects ◽  
Parasitic Infection ◽  
Parasite Burden ◽  
Acute Stage ◽  
Disease Tolerance ◽  
Chronic Stage ◽  
Spatially Resolved

AbstractChagas disease (CD) is a parasitic infection caused by Trypanosoma cruzi protozoa. Over 8 million people worldwide are T. cruzi-positive, 20-30% of which will develop cardiomyopathy, megaoesophagus and/or megacolon. The mechanisms leading to gastrointestinal (GI) symptom development are however poorly understood. To address this issue, we systematically characterized the spatial impact of experimental T. cruzi infection on the microbiome and metabolome across the GI tract. The largest microbiota perturbations were observed in the proximal large intestine in both acute and chronic disease, with chronic-stage effects also observed in the cecum. Strikingly, metabolomic impact of acute-to-chronic stage transition differed depending on the organ, with persistent large-scale effects of infection primarily in the oesophagus and large intestine, providing a potential mechanism for GI pathology tropism in CD. Infection particularly affected acylcarnitine and lipid metabolism. Building on these observations, treatment of infected mice with carnitine-supplemented drinking water prevented acute-stage mortality with no changes in parasite burden. Overall, these results identified a new mechanism of disease tolerance in CD, with potential for the development of new therapeutic regimens. More broadly, these results highlight the potential of spatially-resolved metabolomic approaches to provide insight into disease pathogenesis, with translational applications for infectious disease drug development.

Data Mining in Protein Identification by Tandem Mass Spectrometry

2011 ◽  
pp. 472-478
Author(s):  
Haipeng Wang
Keyword(s):  
Mass Spectrometry ◽  
Data Mining ◽  
Tandem Mass Spectrometry ◽  
Large Scale ◽  
Protein Identification ◽  
Peptide Fragmentation ◽  
Tandem Mass ◽  
Fragmentation Pattern ◽  
Post Translational Modification ◽  
Challenges And Opportunities

Protein identification (sequencing) by tandem mass spectrometry is a fundamental technique for proteomics which studies structures and functions of proteins in large scale and acts as a complement to genomics. Analysis and interpretation of vast amounts of spectral data generated in proteomics experiments present unprecedented challenges and opportunities for data mining in areas such as data preprocessing, peptide-spectrum matching, results validation, peptide fragmentation pattern discovery and modeling, and post-translational modification (PTM) analysis. This article introduces the basic concepts and terms of protein identification and briefly reviews the state-of-the-art relevant data mining applications. It also outlines challenges and future potential hot spots in this field.

scholarly journals A panchromatic spatially resolved study of the inner 500 pc of NGC 1052 – II. Gas excitation and kinematics

2019 ◽  
Vol 489 (4) ◽  
pp. 5653-5668 ◽  
Author(s):  
L G Dahmer-Hahn ◽  
R Riffel ◽  
T V Ricci ◽  
J E Steiner ◽  
T Storchi-Bergmann ◽  
...  
Keyword(s):  
Large Scale ◽  
Near Infrared ◽  
Elliptical Galaxy ◽  
Field Of View ◽  
Spatially Resolved ◽  
Broad Component ◽  
Single Mechanism ◽  
Line Region ◽  
Nir Emission ◽  
Integral Field

ABSTRACT We map the optical and near-infrared (NIR) emission-line flux distributions and kinematics of the inner 320 × 535 pc2 of the elliptical galaxy NGC 1052. The integral field spectra were obtained with the Gemini Telescope using the GMOS-IFU and NIFS instruments, with angular resolutions of 0.88 and 0.1 arcsec in the optical and NIR, respectively. We detect five kinematic components: (1) and (2) two spatially unresolved components: a broad-line region visible in H α, with a full width at half-maximum (FWHM) of ∼3200 km s−1, and an intermediate broad component seen in the [O iii] λλ4959,5007 doublet; (3) an extended intermediate-width component with 280 km s−1 < FWHM < 450 km s−1 and centroid velocities up to 400 km s−1, which dominates the flux in our data, attributed either to a bipolar outflow related to the jets, rotation in an eccentric disc or to a combination of a disc and large-scale gas bubbles; (4) and (5) two narrow (FWHM < 150 km s−1) components, one visible in [O iii], and another visible in the other emission lines, extending beyond the field of view of our data, which is attributed to large-scale shocks. Our results suggest that the ionization within the observed field of view cannot be explained by a single mechanism, with photoionization being the dominant mechanism in the nucleus with a combination of shocks and photoionization responsible for the extended ionization.

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