Photo-clickable microRNA for in situ fluorescence labeling and imaging of microRNA in living cells

2017 ◽  
Vol 53 (48) ◽  
pp. 6452-6455 ◽  
Author(s):  
Lei Huang ◽  
Yingjie Chen ◽  
Lei Chen ◽  
Xiao Xiao ◽  
Xingxing Wang ◽  
...  
Keyword(s):  
Living Cells ◽  
Fluorescence Labeling ◽  
Spatiotemporal Resolution

A photo-clickable microRNA was constructed for in situ fluorescence labeling and imaging of microRNA in living cells with spatiotemporal resolution.

scholarly journals Protein-AMPylierungs-Identifikation in lebenden Zellen

BIOspektrum ◽  
2020 ◽  
Vol 26 (7) ◽  
pp. 743-746
Author(s):  
Tobias Becker ◽  
Pavel Kielkowski
Keyword(s):  
Endoplasmic Reticulum ◽  
Endoplasmic Reticulum Stress ◽  
Neural Development ◽  
Click Reaction ◽  
Living Cells ◽  
Fluorescence Labeling ◽  
Post Translational Modification ◽  
Design Synthesis ◽  
Stress Regulation

AbstractProtein AMPylation is a prevalent protein post-translational modification in human cells involved in endoplasmic reticulum stress regulation and neural development. In this article we describe the design, synthesis and application of a pronucleotide probe suitable for in situ fluorescence imaging and chemical protemics profiling of AMPylated proteins. Our probe utilizes straightforward strain-promoted azidealkyne click reaction for fluorescence labeling in living cells.

DNA nanolantern-based split aptamer probes for in-situ ATP imaging in living cells and lighting up mitochondria

The Analyst ◽  
2021 ◽  
Author(s):  
Yaxin Wang ◽  
Dong-Xia Wang ◽  
Jia-Yi Ma ◽  
Jing Wang ◽  
Yichen Du ◽  
...  
Keyword(s):  
Adenosine Triphosphate ◽  
Cell Metabolism ◽  
Living Cells ◽  
Expression Level ◽  
Specific Analysis

Accurate and specific analysis of adenosine triphosphate (ATP) expression level in living cells can provide valuable information for understanding cell metabolism, physiological activities and pathologic mechanism. Herein, DNA nanolantern-based split...

scholarly journals Mapping High Spatiotemporal-Resolution Soil Moisture by Upscaling Sparse Ground-Based Observations Using a Bayesian Linear Regression Method for Comparison with Microwave Remotely Sensed Soil Moisture Products

2021 ◽  
Vol 13 (2) ◽  
pp. 228
Author(s):  
Jian Kang ◽  
Rui Jin ◽  
Xin Li ◽  
Yang Zhang
Keyword(s):  
Soil Moisture ◽  
Large Scale ◽  
Spatial Scales ◽  
Microwave Remote Sensing ◽  
In Situ Measurements ◽  
Spatial Density ◽  
Linear Regression Method ◽  
Spatiotemporal Resolution ◽  
Pixel Scale

In recent decades, microwave remote sensing (RS) has been used to measure soil moisture (SM). Long-term and large-scale RS SM datasets derived from various microwave sensors have been used in environmental fields. Understanding the accuracies of RS SM products is essential for their proper applications. However, due to the mismatched spatial scale between the ground-based and RS observations, the truth at the pixel scale may not be accurately represented by ground-based observations, especially when the spatial density of in situ measurements is low. Because ground-based observations are often sparsely distributed, temporal upscaling was adopted to transform a few in situ measurements into SM values at a pixel scale of 1 km by introducing the temperature vegetation dryness index (TVDI) related to SM. The upscaled SM showed high consistency with in situ SM observations and could accurately capture rainfall events. The upscaled SM was considered as the reference data to evaluate RS SM products at different spatial scales. In regard to the validation results, in addition to the correlation coefficient (R) of the Soil Moisture Active Passive (SMAP) SM being slightly lower than that of the Climate Change Initiative (CCI) SM, SMAP had the best performance in terms of the root-mean-square error (RMSE), unbiased RMSE and bias, followed by the CCI. The Soil Moisture and Ocean Salinity (SMOS) products were in worse agreement with the upscaled SM and were inferior to the R value of the X-band SM of the Advanced Microwave Scanning Radiometer 2 (AMSR2). In conclusion, in the study area, the SMAP and CCI SM are more reliable, although both products were underestimated by 0.060 cm3 cm−3 and 0.077 cm3 cm−3, respectively. If the biases are corrected, then the improved SMAP with an RMSE of 0.043 cm3 cm−3 and the CCI with an RMSE of 0.039 cm3 cm−3 will hopefully reach the application requirement for an accuracy with an RMSE less than 0.040 cm3 cm−3.

scholarly journals Localization of ‘Candidatus Liberibacter solanacearum’ and Evidence for Surface Appendages in the Potato Psyllid Vector

2016 ◽  
Vol 106 (2) ◽  
pp. 142-154 ◽  
Author(s):  
J. M. Cicero ◽  
T. W. Fisher ◽  
J. K. Brown
Keyword(s):  
Electron Microscopy ◽  
Light And Electron Microscopy ◽  
Fluorescence Labeling ◽  
Potato Psyllid ◽  
Bactericera Cockerelli ◽  
Visual Identification ◽  
Transmission Electron ◽  
Candidatus Liberibacter ◽  
Identification Of Bacteria

The potato psyllid Bactericera cockerelli is implicated as the vector of the causal agent of zebra chip of potato and vein-greening of tomato diseases. Until now, visual identification of bacteria in the genus ‘Candidatus Liberibacter’ has relied on direct imaging by light and electron microscopy without labeling, or with whole-organ fluorescence labeling only. In this study, aldehyde fixative followed by a coagulant fixative, was used to process adult psyllids for transmission electron microscopy (TEM) colloidal gold in situ hybridization experiments. Results indicated that ‘Ca. Liberibacter solanacearum’ (CLso)-specific DNA probes annealed to a bacterium that formed extensive, monocultural biofilms on gut, salivary gland, and oral region tissues, confirming that it is one morphotype of potentially others, that is rod-shaped, approximately 2.5 µm in diameter and of variable length, and has a rough, granular cytosol. In addition, CLso, prepared from shredded midguts, and negatively stained for TEM, possessed pili- and flagella-like surface appendages. Genes implicating coding capacity for both types of surface structures are encoded in the CLso genome sequence. Neither type was seen for CLso associated with biofilms within or on digestive organs, suggesting that their production is stimulated only in certain environments, putatively, in the gut during adhesion leading to multiplication, and in hemolymph to afford systemic invasion.

Optobiochemistry: Genetically Encoded Control of Protein Activity by Light

2021 ◽  
Vol 90 (1) ◽  
Author(s):  
Jihye Seong ◽  
Michael Z. Lin
Keyword(s):  
Protein Function ◽  
Living Cells ◽  
Optical Methods ◽  
Annual Review ◽  
Publication Date ◽  
Protein Activity ◽  
Spatiotemporal Resolution ◽  
Protein Functions ◽  
Protein Classes ◽  
Control Protein

Optobiochemical control of protein activities allows the investigation of protein functions in living cells with high spatiotemporal resolution. Over the last two decades, numerous natural photosensory domains have been characterized and synthetic domains engineered and assembled into photoregulatory systems to control protein function with light.Here, we review the field of optobiochemistry, categorizing photosensory domains by chromophore, describing photoregulatory systems by mechanism of action, and discussing protein classes frequently investigated using optical methods. We also present examples of how spatial or temporal control of proteins in living cells has provided new insights not possible with traditional biochemical or cell biological techniques. Expected final online publication date for the Annual Review of Biochemistry, Volume 90 is June 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

scholarly journals Dynamic nanoscale morphology of the ER surveyed by STED microscopy

2018 ◽  
Vol 218 (1) ◽  
pp. 83-96 ◽  
Author(s):  
Lena K. Schroeder ◽  
Andrew E.S. Barentine ◽  
Holly Merta ◽  
Sarah Schweighofer ◽  
Yongdeng Zhang ◽  
...  
Keyword(s):  
Living Cells ◽  
Stimulated Emission ◽  
Membrane Structures ◽  
Spatiotemporal Resolution ◽  
Sted Microscopy ◽  
Superresolution Microscopy ◽  
Structure And Dynamics ◽  
First Time ◽  
Nanoscale Dynamics ◽  
Conventional Light Microscopy

The endoplasmic reticulum (ER) is composed of interconnected membrane sheets and tubules. Superresolution microscopy recently revealed densely packed, rapidly moving ER tubules mistaken for sheets by conventional light microscopy, highlighting the importance of revisiting classical views of ER structure with high spatiotemporal resolution in living cells. In this study, we use live-cell stimulated emission depletion (STED) microscopy to survey the architecture of the ER at 50-nm resolution. We determine the nanoscale dimensions of ER tubules and sheets for the first time in living cells. We demonstrate that ER sheets contain highly dynamic, subdiffraction-sized holes, which we call nanoholes, that coexist with uniform sheet regions. Reticulon family members localize to curved edges of holes within sheets and are required for their formation. The luminal tether Climp63 and microtubule cytoskeleton modulate their nanoscale dynamics and organization. Thus, by providing the first quantitative analysis of ER membrane structure and dynamics at the nanoscale, our work reveals that the ER in living cells is not limited to uniform sheets and tubules; instead, we suggest the ER contains a continuum of membrane structures that includes dynamic nanoholes in sheets as well as clustered tubules.

scholarly journals Super-Resolution Imaging by Dual Iterative Structured Illumination Microscopy

2021 ◽  
Author(s):  
Anna Loeschberger ◽  
Yauheni Novikau ◽  
Ralf Netz ◽  
Marie-Christin Spindler ◽  
Ricardo Benavente ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Brain Slices ◽  
Super Resolution ◽  
Reconstruction Algorithm ◽  
Living Cells ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Spatiotemporal Resolution ◽  
Coated Pits ◽  
Resolution Imaging

Three-dimensional (3D) multicolor super-resolution imaging in the 50-100 nm range in fixed and living cells remains challenging. We extend the resolution of structured illumination microscopy (SIM) by an improved nonlinear iterative reconstruction algorithm that enables 3D multicolor imaging with improved spatiotemporal resolution at low illumination intensities. We demonstrate the performance of dual iterative SIM (diSIM) imaging cellular structures in fixed cells including synaptonemal complexes, clathrin coated pits and the actin cytoskeleton with lateral resolutions of 60-100 nm with standard fluorophores. Furthermore, we visualize dendritic spines in 70 micrometer thick brain slices with an axial resolution < 200 nm. Finally, we image dynamics of the endoplasmatic reticulum and microtubules in living cells with up to 255 frames/s.

scholarly journals Transport and localization of exogenous myelin basic protein mRNA microinjected into oligodendrocytes.

1993 ◽  
Vol 123 (2) ◽  
pp. 431-441 ◽  
Author(s):  
K Ainger ◽  
D Avossa ◽  
F Morgan ◽  
S J Hill ◽  
C Barry ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Living Cells ◽  
Branch Points ◽  
Rna Granules ◽  
Intracellular Movement ◽  
Directional Movement ◽  
Ionic Detergent ◽  
Mrna Granules

We have studied transport and localization of MBP mRNA in oligodendrocytes in culture by microinjecting labeled mRNA into living cells and analyzing the intracellular distribution of the injected RNA by confocal microscopy. Injected mRNA initially appears dispersed in the perikaryon. Within minutes, the RNA forms granules which, in the case of MBP mRNA, are transported down the processes to the periphery of the cell where the distribution again becomes dispersed. In situ hybridization shows that endogenous MBP mRNA in oligodendrocytes also appears as granules in the perikaryon and processes and dispersed in the peripheral membranes. The granules are not released by extraction with non-ionic detergent, indicating that they are associated with the cytoskeletal matrix. Three dimensional visualization indicates that MBP mRNA granules are often aligned in tracks along microtubules traversing the cytoplasm and processes. Several distinct patterns of granule movement are observed. Granules in the processes undergo sustained directional movement with a velocity of approximately 0.2 micron/s. Granules at branch points undergo oscillatory motion with a mean displacement of 0.1 micron/s. Granules in the periphery of the cell circulate randomly with a mean displacement of approximately 1 micron/s. The results are discussed in terms of a multi-step pathway for transport and localization of MBP mRNA in oligodendrocytes. This work represents the first characterization of intracellular movement of mRNA in living cells, and the first description of the role of RNA granules in transport and localization of mRNA in cells.

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