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13 protocols using expression 11000xl scanner

1

Evaluation of Colony Complexity

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Clones were pinned to 3 replicate LD omni trays, incubated for 7 days, and imaged on an EPSON Expression 11000 XL scanner. Colonies were scored for complexity using the scale in Figure 2.
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2

Characterizing Carbon Ion Beam Profiles

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We used GAF chromic EBT3 films (Ashland Specialty Ingredients GP, NJ USA; Lot # 04022001, Exp. Date: April 2021) for determining the virtual source position of 400 MeV/n carbon ion beam in this study. The film processing and dose profile measurements followed the international protocols [10 (link)]. A preexposure technique was used for the calibration curve derivation [11 (link)]. This was performed by giving each film a priming dose of 2 Gy to homogenize the film density using WHICH facility with a dose of 1 Gy at a carbon ion energy of 400 MeV/u. We then measured the dose homogeneity using a densitometer. Graded doses of 5, 10, 15, 40, 60, 80, 100, 150, and 200 cGy were given to the GAF chromic film to obtain the Hurter-Driffield calibration curve (H-D curve).
All exposed films of depth dose curve were then scanned with an Epson Expression 11000XL scanner in the 48-bit RGB mode (16 bits per color), and the data were saved as tagged image file format (TIFF) and analyzed by the VeriSoft imaging procession software. A red filter was placed on top of the GAF films before scanning to increase the slope of the H-D curve, thereby raising the resolution of the dose-OD curves [12 (link)].
The field size derived from dose 50% of the dose profile at isocenter was then compared to upstream and downstream films with a gap of -15 cm and +15 cm for determining the virtual source position.
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3

Root Morphology Analysis Protocol

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After the above-ground plant parts were removed, root systems and soil media were gently removed from the PVC pots and washed with a gentle stream of water over a wire mesh sieve to remove soil media until the roots were clean. Individual root systems were floated in a 400 × 300 cm acrylic tray filled with 5 mm water. Once placed on the trays, the roots were carefully untangled using plastic forceps to minimize roots’ overlap to ensure quality imagery. An Epson Expression 11000XL scanner captured root morphology images at a 800 dpi resolution (Epson America, Inc., Long Beach, CA, USA). These images were analyzed by WinRHIZO Pro 2009C software (Regent Instruments, Inc., Québec, QC, Canada). The digitized output from the analysis quantified the multiple root growth and development parameters for each plant: root tips (RT, no plant−1), root forks (RF, no plant−1), total root length (TRL, cm plant−1), root surface area (RSA, cm2 plant−1), and root volume (RV, cm3 plant −1)
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4

Root System Analysis Using EZ-Rhizo

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Nine days after seedling transfer, the MS plates were scanned using an EPSON Expression 11000XL scanner in color at 600 dots per inch (dpi) resolution. Scanned images were analyzed using the EZ-Rhizo II software (Armengaud et al., 2009 (link)). Seedlings that were damaged during transfer were excluded from the analysis. The total root length was calculated for each plant by summing the PR and LR lengths. The average LR length was calculated by dividing the total LR length by the number of LRs, and the number of LR per cm was calculated by dividing the total number of LR by the PR length (Gruber et al., 2013 (link)).
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5

Quantitative Protein Expression Profiling

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Each lane of the prepared SDS-PAGE gel was added with 10μl degeneration protein of HEK293T cells and then the gel was placed in a prepared coomassie brilliant blue dye with low velocity for 1 h at room temperature. Then the gel was eluted with destaining solution for 10 min at three times and scanned (Epson Expression 11000XL Scanner). The different expression protein sites of IgG and FBX8 in the gel were cut and placed into special EP tube. The results of the protein sites were analyzed by mass spectrometry.
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6

Tumor Motion Correction for Film Dosimetry

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EBT3 film was used in conjunction with the SNC Patient Film Analysis software (V6.6, Sun Nuclear Corporation) and an Epson Expression 11000XL scanner. As the tumor lesion moves inside the PTV on the TPS a reference frame correction was applied to each of the TPS Dose Planes. However, for the film measurement the film was fixed relative to the motion of the cork insert, and therefore the frame of reference coincides with the position of the tumor. A time‐weighted average correction was applied to each dose value in the TPS dose plane. As the motion of the GTV was sinusoidal with a fixed amplitude and period, the dose values only needed to be corrected in the direction of motion.
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7

Visualizing Bacterial Colony Morphology

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Ten microliters of liquid subcultures was spotted onto 60 mL of colony morphology medium (1% tryptone, 1% agar containing 40 µg/mL Congo red dye and 20 µg/mL Coomassie blue dye) in 100 mm × 100 mm × 15 mm square Petri dishes (LDP D210-16)27 (link). Plates were incubated for up to 5 days in the dark at 25 °C with >90% humidity (Percival CU-22L) and imaged daily with an Epson Expression 11000XL scanner. Images shown are representative of at least six biological replicates from three independent experiments.
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8

Characterization of Natural Yeast Biofilm Diversity

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A total of 78 strains representing a variety of niches and geographical locations from public and private collections [38 (link)–40 (link)] were used for analysis (electronic supplementary material, table S1). All phenotyping was done with homothallic diploid strains, as S. cerevisiae probably exists as a diploid in natural settings. Biofilms were inoculated with overnight YPD cultures (1% yeast extract, 2% peptone, 2% dextrose); four replicates were each scored by two researchers. Mats were inoculated with 2 µl on 0.3% agar low dextrose (0.1%) YPD (LD) 35 × 10 mm plates, sealed and incubated upright at 25°C for 10 days, then imaged and scored for complexity using a four-point scale (electronic supplementary material, figure S1) [17 (link)]. Complex colonies were inoculated using a 96-pin multi-blot replicator on OmniTrays containing 2% agar LD medium. Plates were sealed and incubated at 30°C for 6 days before imaging on an Epson Expression 11000 XL scanner and scored for complexity using a five-point scale (electronic supplementary material, figure S1) [41 (link)].
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9

Measuring Plant Root Morphology

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In both hydroponic experiments, the plant root morphological parameters such as root length (m) and root volume (cm3) of the plants were measured by using a special image analysis software program WinRHIZO (Win/Mac RHIZO Pro V. 2002c Regent Instruments Inc., Québec, QC G1V 1V4, Canada) in combination with Epson Expression 11000XL scanner. From each harvested fresh root samples almost 5.0 g sub-samples were taken. The samples were each (one after the other) placed in the scanner’s tray. Water was added and with the aid of a plastic forceps, the roots were homogenously spread across the tray; and the scanning and analysis done from the WinRhizo system’s interface on a computer connected to the scanner. The total plant root length and volume was then determined as the ratio of sampled root fresh weight to the total root fresh weight.
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10

Yeast Growth Assay Protocols

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For the 100-genomes strains, YPD cultures were grown to saturation (∼24 hr) in a 96-well plate and ∼2 μl per well was transferred to OmniTrays (Nunc 264728) using a 96-pin multi-blot replicator (V&P Scientific no. VP408FP6). For a given assay, a single 96-well plate was pinned to four replicates of three different media types (SLAD, SLAD + PheOH, SLAD + TrpOH). OmniTrays were wrapped with parafilm to prevent drying and incubated at 30C for one week. After incubation, trays were scanned on an Epson Expression 11000 XL scanner, which produced RGB color images with 1200 dpi. For the F5 mapping population, the same procedure was implemented for the 360 segregrants, but only SLAD + PheOH medium was used and with only two replicates per assay. For both the 100-genomes panel and the mapping population, the entire assay was repeated three times.
Follow-up experiments required streaking freezer cultures onto YPD agar, then streaking isolated colonies onto SLAD agar (+ PheOH or TrpOH, when appropriate) and incubating at 30C for 5 days before imaging.
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