mcardinal h2b c 10  (Addgene inc)

Journal: Cell

Article Title: Deciphering cell signaling networks with massively multiplexed biosensor barcoding

doi: 10.1016/j.cell.2021.11.005

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: The following plasmids were used as templates for the fluorescent proteins: H2B-TagRFP (Addgene #99271, a gift from Philipp Keller), mCardinal-H2B-C-10 (Addgene #56162) , piRFP702-N1 (Addgene #45456) , EBFP2-Nucleus-7 (Addgene #55249, a gift from Michael Davidson), and pmCherry-NLS (Addgene #39319) ( ).

Techniques: Recombinant, Plasmid Preparation, Software, Microscopy

Journal: bioRxiv

Article Title: Mechanical competition alters the cellular interpretation of an endogenous genetic programme

doi: 10.1101/2020.10.15.333963

Figure Lengend Snippet: (A) Genes expressed ventrally at the onset of gastrulation. Top: Diagram of a cross-section through an embryo at the beginning of gastrulation. Mesodermal nuclei expressing Snail: blue, mesectodermal nuclei with single-minded: red. Bottom: Schematic of gene expression levels. Twist (black) and Snail (blue) regulate the genes that control shape changes ( fog, T48, mist ). (B) Section of an embryo stained for beta-catenin/armadillo to visualize adherens junctions (pink) and myosin (blue). Junctions in the central (cm) and lateral (lm) mesoderm are apical, the mesectodermal (me) cell has one on apical and one subapical junction, ectodermal junctions are subapical . (C, −D) Cross-sectional views at two time points from a MuVi-SPIM recording of an embryo expressing GAP43::mCardinal (membrane). Pink dots: mesectoderm. timeseries in (C’, D’) Apical surface ‘peels’ with colour-coded apical cell areas. Mesectoderm: white dots. (C”, D”) Apical area from C’ and D’ plotted against cell position (0° is the ventral midline). Each dot represents one cell. Colour-code for rows 7, 8 as in E, mesectoderm magenta. (E) Ventro-lateral views of a confocal recording of an embryo expressing Spider::GFP (white) and sqh::mCherry (green) at a confocal Z-plane 3μm below the surface (Suppl. Movie 2). (E’) Cells were segmented using Spider::GFP and assigned to colour-coded rows. (F-H) Apical areas, total myosin intensity and myosin concentration plotted per row against time (mean and standard deviation). Tracks for ventral rows stop early because the cells are lost from the imaging plane. (I-I’) Example of a lateral mesodermal cell at four time-points in an embryo expressing utrABD::GFP (subapical for cell outlines in I, green; apical in I’; white) and sqh::mCherry (magenta) during formation of a myosin focus. Arrow: local cortical deformation.

Article Snippet: The mCardinal coding sequence was amplified from mCardinal-H2B-C-10 (Addgene plasmid #56162) using a forward primer with the sequence encoding the first 20 amino acids of the GAP43 protein from Bos taurus ( ).

Techniques: Expressing, Gene Expression, Control, Staining, Membrane, Concentration Assay, Standard Deviation, Imaging

Journal: bioRxiv

Article Title: Mechanical competition alters the cellular interpretation of an endogenous genetic programme

doi: 10.1101/2020.10.15.333963

Figure Lengend Snippet: (A) Embryos were imaged using Multi-View SPIM. The resulting data sets were fused into a single 3D stack for each timepoint. Ventral side top left. (B, C) Apical (outside) and basal surface masks were defined semi-automatically. These masks were then used to extract the apical surface of the embryo . (D, D’) Maximum intensity projections along the apical-basal (D) and anterior-posterior (D’) direction of an embryo co-expressing GAP43::mCherry, Snail::MS2 and MCP::mCherry 4 min before the initiation of ventral furrow formation. The white spots represent sites of snail::MS2 RNA in the nuclei of mesodermal cells. Yellow and blue dots mark the positions of the adjacent mesectodermal cell rows. (E-I) Confocal Z-plane 2μm from the surface and Z-sections (E’-I’) over the course of furrow invagination. The mesectodermal cell rows meet at the midline. Back-tracing from this time point can be used to determine the edge of the mesoderm in unmarked embryos. (J, J’) Surface peels extracted from MuVi SPIM images at −2 and 6 min from initiation of ventral furrow formation. White dots indicate the mesectodermal cells as determined by backtracing. Cell rows are colour-coded with numbering coordinated operationally around row 6, which is the last non-stretching row and easily identifiable in all movies, regardless of imaging angle. The width of the mesoderm varies along the anterior-posterior axis, with a width of less than 18 cells in some areas. (K-O) Each panel is from one time-point from a MuVi SPIM recording of an embryo expressing GAP43::mCardinal, with three of the three images showing first, cross-sectional views; secondly, apical surface ‘peels’ extracted from the ventral half of the central one third of the embryo, And finally and third, the apical areas plotted against cell position along the left-right axis (the centre, 0°, is the ventral midline of the embryo). Each dot represents one cell. Apical cell areas measured from segmented images were colour-coded and overlayed on the original image. Mesectodermal cells are marked as white dots in the surface peels and as magenta dots in the plots. For the description in this figure, we define t= 0 min as the time when cells in the central four rows have constricted on average by at least 20%.

Article Snippet: The mCardinal coding sequence was amplified from mCardinal-H2B-C-10 (Addgene plasmid #56162) using a forward primer with the sequence encoding the first 20 amino acids of the GAP43 protein from Bos taurus ( ).

Techniques: Expressing, Imaging