Branches of similar size (2 cm in length) were sampled from colonies in the pH 7.2 and 8.0 treatments and placed in a 10% NaClO solution for 2 hours to remove tissues. Skeletons were rinsed several times in tap water, followed by ultrapure H2O, and then dried at 40°C for at least 24 hours. Samples (five replicates per pH condition) were coated with gold/palladium and observed at 4 kV with a JEOL 6010LV electron microscope. Diameters of the corallite calyxes were measured using manufacturer-provided SMILE VIEW software (JEOL). A total of 181 measurements were performed across 10 fragments (data S5B).
Smile view software
Manufactured by JEOL
Sourced in Japan
SMILE VIEW software is a data processing and analysis tool developed by JEOL. It provides functionalities for visualizing and analyzing data generated from various JEOL laboratory equipment.
Automatically generated - may contain errors
Lab products found in correlation
Epibrassinolide, by Merck Group (1 mentions)
Superfrost microscope slide, by Thermo Fisher Scientific (1 mentions)
Paraformaldehyde (pfa), by Merck Group (1 mentions)
Bioplotter, by EnvisionTEC (1 mentions)
S 3400 sem, by Hitachi (1 mentions)
E102 ion sputter, by Hitachi (1 mentions)
Crossbeam 550, by Zeiss (1 mentions)
Em cpd030, by Leica camera (1 mentions)
Jem 1400 electron microscope, by JEOL (1 mentions)
4 protocols using smile view software
1
Coral Skeleton Morphometry Analysis
2
Investigating Brassinosteroid Regulation of Xylem Formation
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Four-week-old wild-type (WT) Populus “Nanlin 895” were incubated with the hormone solutions. The concentrations for the treatments were 0 (control), 1, 10, and 50 μM BL (Epibrassinolide; Sigma–Aldrich, St. Louis, MO, United States). BL-treated stems were subjected to PdC3H17 expression analysis by quantitative real-time polymerase chain reaction (qRT-PCR). Three biological replicates were performed independently.
To analyze the responsiveness of BRs or auxin with BRs on xylem formation, at least three independent lines of root-removed control and transgenic seedlings were grown on the 1/2 MS medium with or without PCZ/(PCZ + IBA) at various determined concentrations for 21 days (8 h for gene expression analysis), and 0.3-cm segments were taken from their basal stems. These segments were fixed in 4% paraformaldehyde (Sigma-Aldrich) at 4°C for 4 days, dehydrated in graded ethanol series, and embedded into paraplast. The 5-μm sections were obtained using a Leica RM2235 rotary microtome and adhered to Superfrost microscope slides (Thermo). The sections were stained with 0.1% toluidine blue and observed using an Olympus X51 light microscope (Olympus). Radial widths of xylem were measured in three independent replicates using the SmileView software (JEOL).
To analyze the responsiveness of BRs or auxin with BRs on xylem formation, at least three independent lines of root-removed control and transgenic seedlings were grown on the 1/2 MS medium with or without PCZ/(PCZ + IBA) at various determined concentrations for 21 days (8 h for gene expression analysis), and 0.3-cm segments were taken from their basal stems. These segments were fixed in 4% paraformaldehyde (Sigma-Aldrich) at 4°C for 4 days, dehydrated in graded ethanol series, and embedded into paraplast. The 5-μm sections were obtained using a Leica RM2235 rotary microtome and adhered to Superfrost microscope slides (Thermo). The sections were stained with 0.1% toluidine blue and observed using an Olympus X51 light microscope (Olympus). Radial widths of xylem were measured in three independent replicates using the SmileView software (JEOL).
3
3D-Printed PLGA Scaffold Fabrication
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The 3D-printed PLGA scaffolds were created using a 3DP system (Bioplotter; Envision TEC GmbH, Gladbeck, Germany) according to previous studies. The PLGA (molecular weight 50,000 to 75,000; poly lactic acid/poly glycolic acid 50:50) was purchased from Sigma-Aldrich. To fabricate the 3D-printed PLGA scaffolds, the PLGA powder was melted at 135°C and centrifuged at 100 × g for 10 min at ∼140°C. The melted PLGA was dispensed via a 27-gauge metal needle to construct the 3D interconnected scaffolds. The dispensing temperature was 135°C, the pressure was ∼6,000 kPa, and the XY dispense head speed was set at 80 mm/min. The microstructure of the scaffolds was examined by a scanning electron microscopy (SEM). The SMILE view software (JEOL, Tokyo, Japan) was used to determine the size of the pores in the scaffolds from the SEM images.
4
Ultrastructural Analysis of Fungal Hyphae
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For transmission electron microscopy analysis, hyphae were cultured on PDA medium for 18 hr and prefixed with 2% (w/v) glutaraldehyde containing 0.1 M phosphate buffer (pH 7) at 4 °C for 10 hr, followed by staining with 1% osmium tetroxide in phosphate buffer at room temperature for an additional hour. After dehydration, tissues were embedded in epoxy resin. Ultrathin sections were stained with uranyl acetate and lead citrate and visualised under a JEM 1400 electron microscope (JEOL, Japan). The cell wall thickness was measured using SMile View software (JEOL, Japan).
For SEM analysis, hyphae were cultured on PDA medium at 37 °C for 48 hr, and agar blocks containing fungal cells were fixed as described for the transmission electron microscopy analysis. The blocks were then dehydrated by passage through a graded series of ethanol solutions, replaced with isoamyl acetate, and dried using the critical‐point method (EM CPD030; Leica, Germany). After sputter coating with platinum‐palladium (using an E102 Ion sputter; Hitachi, Japan), samples were visualised using an S‐3400 SEM (Hitachi, Japan). The proportion of spike area to the total surface area of each conidium was measured using Image J. For field emission‐scanning electron microscopy analysis, dehydrated samples were visualised on poly‐L‐lysine coated silicon wafers under CrossBeam 550 (Zeiss).
For SEM analysis, hyphae were cultured on PDA medium at 37 °C for 48 hr, and agar blocks containing fungal cells were fixed as described for the transmission electron microscopy analysis. The blocks were then dehydrated by passage through a graded series of ethanol solutions, replaced with isoamyl acetate, and dried using the critical‐point method (EM CPD030; Leica, Germany). After sputter coating with platinum‐palladium (using an E102 Ion sputter; Hitachi, Japan), samples were visualised using an S‐3400 SEM (Hitachi, Japan). The proportion of spike area to the total surface area of each conidium was measured using Image J. For field emission‐scanning electron microscopy analysis, dehydrated samples were visualised on poly‐L‐lysine coated silicon wafers under CrossBeam 550 (Zeiss).
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