Journal: Microbiology Spectrum

Article Title: Imaging Minimal Bacteria at the Nanoscale: a Reliable and Versatile Process to Perform Single-Molecule Localization Microscopy in Mycoplasmas

doi: 10.1128/spectrum.00645-22

Figure Lengend Snippet: PALM imaging of an F-type ATPase in Mycoplasma mycoides subsp. capri . M. mycoides subsp. capri mEos3.2-0575 cells, expressing a mEos-fused variant of the β-subunit of the ATPase F 1-like domain, were imaged by PALM. In this M. mycoides subsp. capri mutant, the fluorescent fusion protein is expressed from the native genomic locus and replaces the wild-type variant. The data presented here correspond to a single representative field of view (512 by 512 pixels; pixel size = 160 nm). (A) Sample images of M. mycoides subsp. capri mEos3.2-0575 cells. For each field of view, the images correspond to epifluorescence (diffraction limited) (top), superresolved reconstruction (40-nm pixel) (middle), and Tesseler segmentation (bottom). Scale bar = 1 μm. (B) Tesseler clustering of the fluorescence signal. The number of clusters per Tesseler-segmented object was computed. The bar graphs display the distribution of the numbers of clusters per object. (C) Object and cluster sizes. The dot plot presents the area (in nm 2 ) of each object and cluster segmented by Tesseler, to which a boxplot showing the median, interquartile range, minimum, and maximum values is overlaid. The median value of each data set is indicated. (D) Evaluation of the PALM imaging pointing accuracy. Inset, example of the tracks computed using PALMTracer from which the MSD 0 and pointing accuracy values are derived. The bar graphs display the distribution of the pointing accuracies derived from each track. The median value of the data set is indicated.

Article Snippet: The PALMTracer plugin for MetaMorph ( ) was used to process image stacks with a specific intensity threshold for each data set, to enable the generation of the localization tables.

Techniques: Imaging, Expressing, Variant Assay, Mutagenesis, Fluorescence, Derivative Assay

Journal: Microbiology Spectrum

Article Title: Imaging Minimal Bacteria at the Nanoscale: a Reliable and Versatile Process to Perform Single-Molecule Localization Microscopy in Mycoplasmas

doi: 10.1128/spectrum.00645-22

Figure Lengend Snippet: PALM imaging of an F-type ATPase in Mycoplasma mycoides subsp. capri . M. mycoides subsp. capri mEos3.2-0575 cells, expressing a mEos-fused variant of the β-subunit of the ATPase F 1-like domain, were imaged by PALM. In this M. mycoides subsp. capri mutant, the fluorescent fusion protein is expressed from the native genomic locus and replaces the wild-type variant. The data presented here correspond to a single representative field of view (512 by 512 pixels; pixel size = 160 nm). (A) Sample images of M. mycoides subsp. capri mEos3.2-0575 cells. For each field of view, the images correspond to epifluorescence (diffraction limited) (top), superresolved reconstruction (40-nm pixel) (middle), and Tesseler segmentation (bottom). Scale bar = 1 μm. (B) Tesseler clustering of the fluorescence signal. The number of clusters per Tesseler-segmented object was computed. The bar graphs display the distribution of the numbers of clusters per object. (C) Object and cluster sizes. The dot plot presents the area (in nm 2 ) of each object and cluster segmented by Tesseler, to which a boxplot showing the median, interquartile range, minimum, and maximum values is overlaid. The median value of each data set is indicated. (D) Evaluation of the PALM imaging pointing accuracy. Inset, example of the tracks computed using PALMTracer from which the MSD 0 and pointing accuracy values are derived. The bar graphs display the distribution of the pointing accuracies derived from each track. The median value of the data set is indicated.

Article Snippet: Based on the acquired image stacks, the localization of each individual fluorescence emitter was determined through the PALMTracer plugin under MetaMorph, and then superresolved images were reconstructed and analyzed by automatic Voronoï-based segmentation of these localizations ( , Fig. S2).

Techniques: Imaging, Expressing, Variant Assay, Mutagenesis, Fluorescence, Derivative Assay

Journal: Nucleic Acids Research

Article Title: A dominant-negative SOX18 mutant disrupts multiple regulatory layers essential to transcription factor activity

doi: 10.1093/nar/gkab820

Figure Lengend Snippet: List of software, algorithms and publicly available datasets that were used in this manuscript

Article Snippet: Here, we used the Metamorph plugin PalmTracer to obtain average diffusion coefficients for the bound and unbound fractions of SOX18 (0.02 μm /s and 0.43 μm /s) and SOX18 RaOp (0.02 and 0.27 μm /s).

Techniques: Software, Microscopy, Fluorescence