Tm 3000 tabletop sem

Whole mount of mature dry seeds from WT Col-0 and the gosamt1-1gosamt1-2 mutant line were attached to double-sided carbon sealing tape and coated with gold in a 108 manual sputter coater (Ted Pella Inc.). Morphological details of the seeds surface were photographed using a scanning electron microscope (Hitachi TM3000 Tabletop SEM) set to 15 kV. A minimum of three biological replicates were used for each genotype.

To locate vessels exposed on the surface of thin sections and to obtain element maps of vessel-filling material and near-vessel bone, we used a Hitachi TM-3000 Tabletop SEM coupled with a Quantax 70 Energy Dispersive X-ray Spectrophotometer (EDS) system (Hitachi High-Tech America, Inc., Schaumburg, IL). In initial analyses, we left the samples uncoated, but surrounded them with aluminum tape to improve conductivity, and used the Quantax70 to interpret and visualize the EDS data. Based on these preliminary analyses, we identified specimen 2018-L2 (longitudinal section of a gastralium fragment, see Table 1) as the best candidate for quantitative EDS spot analysis due to the abundance of vessels exposed on the surface of the section.
We then collected spot analyses from 2018-L2 on an FEI Quanta 600FEG environmental SEM (FEI Company, Hillsboro, OR) with both back scattered electron (BSE) and secondary electron (SE) capability, operating at a voltage of 5–20 kV. The sample was coated in a mixture of gold and palladium and surrounded by aluminum tape to improve conductivity. We used a Bruker EDX to carry out spot analyses of elemental concentrations and to generate additional elemental maps on vessel fills and lacuna fills.

The microstructures of the sintered samples were investigated using two scanning electron microscopes. The first was a Hitachi TM3000—TableTop SEM from Tokyo, Japan with an electron beam energy of 5 keV and 15 keV. The second high-resolution SEM was an FEI Nova NanoSEM 650 (Eindhoven, The Netherlands) with an electron beam energy of 10 keV. The low vacuum mode was used for microstructure observation; therefore, metal sputtering was not used. Energy dispersive mapping analysis was performed for certain samples.

Physical characterization of the samples consisted of roughness and composition measurements. Atomic force microscope topography was used to measure the roughness of each thin film. This was completed using a Dimension Icon AFM (Bruker, Santa Barbara, CA, USA) using the ScanAsyst mode and SCANASYST-AIR cantilevers. A minimum of three 10  $\mathrm{\mu }$ m AFM images (512 pixels  $\times$  512 pixels) were taken of each sample. Nanoscope Analysis software (Bruker, Santa Barbara, CA, USA) was used to find the three-dimensional area of each image. For the NiCo thin films the elemental composition was measured. Scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) measurements were completed using a TM3000 Tabletop SEM (Hitachi, Tokyo, Japan) and a Quantax 70 EDS attachment (Bruker, Madison, WI, USA). Images and EDS data were taken at $\times 60$ magnification, and Quantax 70 software was used to obtain the Ni and Co compositions from the EDS spectra.

For scanning electron microscopy (SEM) observations of the periostracum, shells were cleaned using a fine brush and directly mounted by carbon tape on aluminium stubs. To observe protoconch and teleoconch sculpture, shells were placed in diluted (10%) household bleach for 30 s and cleaned by a fine brush to remove debris and also the periostracum. The cleaned shells were washed for a few times in MilliQ water and twice in 99% ethanol, before being mounted on aluminium stubs. For radula observations, the radula ribbon was dissected from the limpets using tungsten needles and then cleaned in 10% household bleach to dissolve any remaining soft tissue. The cleaned radulae were then washed and mounted in the same manner as shells. All SEM stubs prepared were observed uncoated at 15 kV using a Hitachi TM-3000 tabletop SEM.

Elemental analyses were obtained using a EuroVector EA3000 Elemental Analyser (EuroVector S.p.A., Milan, Italy). FTIR spectra were recorded on a Bruker Vertex 80 (Bruker Optics GmbH & Co. KG, Ettlingen, Germany) as KBr disks. Energy-dispersive X-ray spectroscopy (EDS) was performed on a Hitachi TM3000 TableTop SEM (Hitachi High-Technologies Corporation, Tokyo, Japan) with Bruker QUANTAX 70 EDS equipment. Thermal properties were studied on a Thermo Microbalance TG 209 F1 Iris (NETZSCH) from 25 to 800 °C, at a rate of 10 °C·min⁻¹ in He flow (30 mL/min). The absorption spectra were investigated using an Agilent Cary 60 UV–Vis spectrophotometer (USA). High-resolution electrospray mass spectrometric (HR-ESI-MS) detection was performed at the Center of Collective Use “Mass spectrometric investigations” SB RAS in negative mode within the 500–3000 m/z range on an electrospray ionization quadrupole time-offlight (ESI-q-TOF) high-resolution mass spectrometer Maxis 4G (Bruker Daltonics, Bremen, Germany).