Fast Super-Resolution Imaging with Ultra-High Labeling Density Achieved by Joint Tagging Super-Resolution Optical Fluctuation Imaging (JT-SOFI)

Previous stochastic localization-based super-resolution techniques are largely limited by the labeling density and the fidelity to the morphology of specimen. We report on an optical super-resolution imaging scheme implementing joint tagging using multiple fluorescent blinking dyes associated with super-resolution optical fluctuation imaging (JT-SOFI), achieving ultra-high labeling density super-resolution imaging. To demonstrate the feasibility of JT-SOFI, quantum dots with different emission spectra were jointly labeled to the tubulin in COS7 cells, creating ultra-high density labeling. After analyzing and combining the fluorescence intermittency images emanating from spectrally resolved quantum dots, the microtubule networks are capable of being investigated with high fidelity and remarkably enhanced contrast at sub-diffraction resolution. The spectral separation also significantly decreased the frame number required for SOFI, enabling fast super-resolution microscopy through simultaneous data acquisition. As the joint-tagging scheme can decrease the labeling density in each spectral channel, we can faithfully reflect the continuous microtubule structure with high resolution through collection of only 100 frames per channel. The improved continuity of the microtubule structure is quantitatively validated with image skeletonization, thus demonstrating the advantage of JT-SOFI over other localization-based super-resolution methods.Comment: 19 pages, 4 figures, with S

Multi-modality Super-resolution Optical Imaging of Living System

EI

A Comparison of Linear Conventional and Nonlinear Microbial Models for Simulating Pulse Dynamics of Soil Heterotrophic Respiration in a Semi-Arid Grassland

As a critical process in regulating terrestrial feedback to climate change, soil heterotrophic respiration is commonly simulated with the first-order kinetics in current Earth system models. Compared with the first-order kinetic models, explicit microbial models are expected to better simulate nonlinear carbon (C)-cycle phenomena, such as the pulse dynamics of soil heterotrophic respiration driven by dry-rewetting cycles in grasslands. However, these two types of models (i.e., linear conventional and nonlinear microbial models) have never been compared based on in situ observations of soil heterotrophic respiration in the semi-arid grassland, which is significantly affected by the dry-rewetting events. Here, based on the field data of soil heterotrophic respiration in a semi-arid grassland in northern China, we first showed that the shift from a conventional linear model to a nonlinear microbial model did not substantially improve the simulation of soil heterotrophic respiration. Then, we quantified the contributions of different moisture-response functions to the uncertainty in simulating the soil C dynamics. The results showed that the selection of moisture-response functions combined with parameterization in the soil C models dominated the modeled uncertainties in soil heterotrophic respiration. These findings suggest that both the conventional linear model and nonlinear microbial model can simulate well the pulse dynamic of soil heterotrophic respiration in grasslands with an improved parameterization of water regulation on soil carbon decomposition. This study also calls for more observations of nonlinear C phenomena for reducing the simulation uncertainty on soil C cycling in Earth system models

A Comparison of Linear Conventional and Nonlinear Microbial Models for Simulating Pulse Dynamics of Soil Heterotrophic Respiration in a Semi‐Arid Grassland

As a critical process in regulating terrestrial feedback to climate change, soil heterotrophic respiration is commonly simulated with the first-order kinetics in current Earth system models. Compared with the first-order kinetic models, explicit microbial models are expected to better simulate nonlinear carbon (C)-cycle phenomena, such as the pulse dynamics of soil heterotrophic respiration driven by dry-rewetting cycles in grasslands. However, these two types of models (i.e., linear conventional and nonlinear microbial models) have never been compared based on in situ observations of soil heterotrophic respiration in the semi-arid grassland, which is significantly affected by the dry-rewetting events. Here, based on the field data of soil heterotrophic respiration in a semi-arid grassland in northern China, we first showed that the shift from a conventional linear model to a nonlinear microbial model did not substantially improve the simulation of soil heterotrophic respiration. Then, we quantified the contributions of different moisture-response functions to the uncertainty in simulating the soil C dynamics. The results showed that the selection of moisture-response functions combined with parameterization in the soil C models dominated the modeled uncertainties in soil heterotrophic respiration. These findings suggest that both the conventional linear model and nonlinear microbial model can simulate well the pulse dynamic of soil heterotrophic respiration in grasslands with an improved parameterization of water regulation on soil carbon decomposition. This study also calls for more observations of nonlinear C phenomena for reducing the simulation uncertainty on soil C cycling in Earth system models

Identification of miRNA–mRNA Networks Associated with Pigeon Skeletal Muscle Development and Growth

The growth and development of skeletal muscle determine the productivity of pigeon meat production, and miRNA plays an important role in the growth and development of this type of muscle. However, there are few reports regarding miRNA regulating the growth and development of skeletal muscle in pigeons. To explore the function of miRNA in regulating the growth and development of pigeon skeletal muscle, we used RNA sequencing technology to study the transcriptome of pigeons at two embryonic stages (E8 and E13) and two growth stages (D1 and D10). A total of 32,527 mRNAs were identified in pigeon skeletal muscles, including 14,378 novel mRNAs and 18,149 known mRNAs. A total of 2362 miRNAs were identified, including 1758 known miRNAs and 624 novel miRNAs. In total, 839 differentially expressed miRNAs (DEmiRNAs) and 11,311 differentially expressed mRNAs (DEGs) were identified. STEM clustering analysis assigned DEmiRNAs to 20 profiles, of which 7 were significantly enriched (p-value < 0.05). These seven significantly enriched profiles can be classified into two categories. The first category represents DEmiRNAs continuously downregulated from the developmental stage to the growth stage of pigeon skeletal muscle, and the second category represents DEmiRNAs with low expression at the development and early growth stage, and significant upregulation at the high growth stage. We then constructed an miRNA–mRNA network based on target relationships between DEmiRNAs and DEGs belonging to the seven significantly enriched profiles. Based on the connectivity degree, 20 hub miRNAs responsible for pigeon skeletal muscle development and growth were identified, including cli-miR-20b-5p, miR-130-y, cli-miR-106-5p, cli-miR-181b-5p, miR-1-z, cli-miR-1a-3p, miR-23-y, cli-miR-30d-5p, miR-1-y, etc. The hub miRNAs involved in the miRNA–mRNA regulatory networks and their expression patterns during the development and growth of pigeon skeletal muscle were visualized. GO and KEGG enrichment analysis found potential biological processes and pathways related to muscle growth and development. Our findings expand the knowledge of miRNA expression in pigeons and provide a database for further investigation of the miRNA–mRNA regulatory mechanism underlying pigeon skeletal muscle development and growth

Study on Heat Resistance of Peony Using Photosynthetic Indexes and Rapid Fluorescence Kinetics

To investigate the effects of high-temperature stress on the chlorophyll fluorescence induction kinetics of peony and to determine indicators for the rapid screening of varieties responding to high temperatures, three four-year-old peony variety, ‘Fengdanbai’, ‘Huhong’, and ‘Yinhongqiaodui’, were selected as materials. The photosynthetic curves (Pn-PAR) and fast chlorophyll fluorescence curves (OJIP curves) of peony leaves were measured at different times under high-temperature stress conditions (40 °C), the changes in the photosynthetic characteristics of different peony varieties under high-temperature stress were analyzed, and the heat tolerance of peony was evaluated. The results showed that ‘Huhong’ grew well within 16 days, while all of the other varieties showed obvious wilting at 6–9 days. High temperatures damaged the structure and function of the photosystem of peony leaves, indicating that the maximum net photosynthetic rate (Pnmax), apparent quantum efficiency (AQE), maximum photochemical efficiency (Fv/Fm), and photosynthetic performance index (PIABS) all tended to decrease under high-temperature stress, while the rate of closing the PS II reaction center (Mo) and the absorption per reaction center (ABS/RC), the capture (TRo/RC), and the dissipation (Dio/RC) of light energy per reaction center showed an overall increasing trend. The ability to cope with high-temperature stress differed among varieties, and the heat tolerance was determined to be in the descending order of ‘Fengdanbai’ ABS, Dio/RC, and Mo could be used as indicators of peony tolerance to high-temperature stress. The results of the study can provide a basis for the screening of heat-tolerant peony species and peony heat defense in the Jiangnan area

GMars‑Q Enables Long-Term Live-Cell Parallelized Reversible Saturable Optical Fluorescence Transitions Nanoscopy

The recent development of reversibly switchable fluorescent proteins (RSFPs) has promoted reversible saturable optical fluorescence transitions (RESOLFT) nanoscopy as a general scheme for live-cell super-resolution imaging. However, continuous, long-term live-cell RESOLFT nanoscopy is still hindered mainly because of the unsatisfactory properties of existing RSFPs. In this work, we report GMars-Q, a monomeric RSFP with low residual off-state fluorescence and strong fatigue resistance attributed to a biphasic photobleaching process. We further demonstrate that GMars-Q is particularly suitable for long-term parallelized RESOLFT nanoscopy as it supports an order of magnitude longer imaging durations than existing RSFPs. The excellent photophysical properties of GMars-Q also suggest that it would be of general interest for other RESOLFT nanoscopic methods

STED optical super-resolution microscopy with fluorescent NV-centers

In this work, we present the application of RESOLFT nanoscopy in subcellular organelle imaging. Fluorescent nano-diamond (FND) has been imaged as a photostable inorganic dye, with large stimulated emission cross-section. FND nanoparticles as well as bulk FNDs grown in a large diamond have been studied with our STED system. ? OSA 2013.EI