Dfc7000 t digital camera

Soil samples containing needle nematodes resembling L. leptocephalus were collected at a depth of 10 to 30 cm from the rhizosphere of grapevine in the area of Thessaloniki, northern Greece. Nematodes were extracted from soil by modified sieving and a decanting method (Brown and Boag, 1988 ). Extracted specimens were handpicked, heat killed, fixed in TAF (triethanolamine formalin), processed using glycerol by a slow evaporation method, and mounted on permanent slides (Hooper, 1986 ). The light micrographs and measurements of nematodes population, including the main diagnostic characteristics, were performed using a Leica DM6 compound microscope with a Leica DFC7000 T digital camera. All other abbreviations used were as defined in the study by Jairajpuri and Ahmad (1992) .

Ten sunflower root pieces with O. cumana seedlings attached were cut from the sunflower plants of the rhizotron assay at 14, 21, 28, and 35 dpi using a binocular microscope. Samples were prepared as in Chabaud et al. (2022 ). Half of the samples were fixed in ethanol: acetic acid (3:1 by volume) for 10 min under vacuum, cleared in chloral hydrate 5 g/ml for 48 h under agitation and visualized with an Axioplan 2 light microscope (Zeiss, Jena, Germany). The remaining samples were fixed in FAA solution (10% formaldehyde, 5% acetic acid, and 50% ethanol) for 5 min under vacuum, dehydrated in alcohol series, and embedded in Technovit 7100 resin (Heraeus Kulzer, Germany). Thin sections of 10 µm were then made using a Reichert-Jung 2040 microtome (Leica Biosystems, Nussloch, Germany), stained with 0.2% toluidine blue for 3 min, mounted in DePeX mounting medium and scanned using a NanoZoomer image scanner (Hamamatsu Photonics, Japan). For the detection of phenolic compounds, hand-cut sections (with a razor blade) obtained from fresh root samples at 14, 21, 28, and 35 dpi were observed under epifluorescence (340–380 nm), as described in Lozano-Baena et al. (2007 (link)), in a Leica DM6 compound microscope with a Leica DFC7000 T digital camera (Wetzlar, Germany).

Soil samples were collected at a depth of 20 to 40 cm from the rhizosphere of a grapevine grafted on 1103-Paulsen of the Institute of Plant Breeding and Genetic Resources, Thermi, Thessaloniki, Greece. Nematodes were extracted from soil by modified sieving and decanting method (Brown and Boag, 1988 ). Extracted specimens were heat killed, fixed in TAF, processed to glycerol by a slow evaporation method, and mounted on permanent slides (Hooper, 1986 ). The light micrographs and measurements of nematode populations including the main diagnostic characteristics (i.e., de Man indices, body length, odontostyle length, lip region, tail shape, amphid shape, and oral aperture-guiding ring) were performed using a Leica DM6 compound microscope with a Leica DFC7000 T digital camera. All abbreviations were used as defined in Jairajpuri and Ahmad (1992) .

Tibias isolated from 3-month-old male mice were fixed in 4% paraformaldehyde and then decalcified with 10% EDTA. The decalcified bones were embedded in paraffin and sectioned. Sections were deparaffinized and hydrated through xylene and graded concentrations of alcohol. Antigen retrieval was performed with hyaluronidase. Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100). Slides were then incubated with secondary antibodies against rabbit IgG [Alexa Fluor 568 (A10042, Thermo Fisher Scientific; 1:500)] and goat IgG [Alexa Fluor 488 (A11055, Thermo Fisher Scientific; 1:500)], followed by DAPI staining. Images were acquired with a DFC7000 T digital camera (Leica, Wetzlar, Germany) under a DM4B microscope (Leica).

To observe the localization of MoRPD3, a translational fusion construct of MoRPD3 and GFP (expressed under the native promoter of the MoRPD3 gene) was prepared and introduced into the wild-type strain and a strain expressing histone H1-RFP. For observation of the fluorescence signal in hyphae, a sterilized glass slide was immersed and coated with complete medium agar and then inoculated with mycelia of the strain expressing MoRPD3-GFP. Following incubation at 25°C for 5 days, nuclei were stained using the Hoechst 33342 dye (Invitrogen, Waltham, MA, USA) according to the manufacturer’s instructions prior to observation of MoRPD3-GFP localization in mycelia. In spores, the MoRPD3-GFP/H1-RFP strain was used to examine the localization of MoRPD3, compared to its localization in fungal nuclei. Microscopy was performed with a Leica DM2500 light microscope, and images were taken with a Leica DFC7000 T digital camera. Excitations were 340 to 380 nm for Hoechst 33342 dye, 480/40 nm for GFP, and 515 to 560 nm for RFP. Images were processed using LAS X software.

Digital images of nymphs, adults, and isolated adult wings were taken using a DFC7000 T digital camera (Leica, Wetzlar, Germany) connected to an M165 FC stereomicroscope (Leica). Signal intensity, measured as the mean grayscale value on 8-bit images, was determined using the image processing software ImageJ Fiji (https://fiji.sc).