Soil organic matter was determined using the K2Cr2O7 titration method. Soil available P was extracted with 0.5 M NaHCO3 (pH 8.5) and then determined using the Olsen method [37 ], and soil available K was extracted with 1 M ammonium acetate (pH 7.0), and measured by atomic absorption spectrometry (NovAA300, Analytik Jena AG). The total N content and the atom% 15N of soils and plants were determined using a stable isotope ratio mass spectrometer (Isoprime100 and Vario Pyrd/Cube, Elementar, Germany). Soil mineral N content (Nmin: NH4+-N and NO3–-N) was extracted with 1M KCl and determined using a flow injection analyzer (FLA star 5000 Analyzer, Foss, Denmark). Soil microbial biomass N content (MBN) was determined by chloroform fumigation-extraction protocol, extracted with 0.5M K2SO4, and analyzed using a multi N/C analyzer (Multi N/C 3100/HT1300, Analytik Jena AG, Germany). The atom% 15N of Nmin and MBN was determined using a stable isotope ratio mass spectrometer (Isoprime100 and Vario Pyrd/Cube, Elementar, Germany).
Isoprime 100
Manufactured by Elementar
Sourced in Germany, United Kingdom
The Isoprime 100 is a stable isotope ratio mass spectrometer (IRMS) designed for high-precision measurement of the isotopic composition of various elements, such as carbon, nitrogen, oxygen, and hydrogen. It is a versatile and reliable instrument that can be used in a range of applications, including environmental research, agricultural studies, and forensic analysis.
Automatically generated - may contain errors
Lab products found in correlation
Vario pyro cube, by Elementar (4 mentions)
Vario isotope select, by Elementar (2 mentions)
Flash 2000 elemental analyzer, by Thermo Fisher Scientific (1 mentions)
Chns elemental analyzer, by Elementar (1 mentions)
Vario micro cube, by Elementar (1 mentions)
Poraplot q, by Agilent Technologies (1 mentions)
Delta 5 advantage, by Thermo Fisher Scientific (1 mentions)
Flash ea 1112, by Thermo Fisher Scientific (1 mentions)
Gc 14a, by Shimadzu (1 mentions)
Asap 2020, by Micromeritics (1 mentions)
Multi n c 3100 ht1300, by Analytik Jena (1 mentions)
Novaa300, by Analytik Jena (1 mentions)
Vario pyrd cube, by Elementar (1 mentions)
Fla star 5000 analyzer, by Foss (1 mentions)
Euro ea3028, by Eurovector (1 mentions)
Isotope cube, by Elementar (1 mentions)
K15no3, by Merck Group (1 mentions)
Vario el cube, by Elementar (1 mentions)
Uv5500, by Metash (1 mentions)
13c sucrose, by Merck Group (1 mentions)
32 protocols using isoprime 100
1
Soil Nutrient Analyses Using Standard Methods
2
Soil-Plant Carbon-Nitrogen-Phosphorus Dynamics
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Total carbon and nitrogen concentration of plant and soil samples were determined sequentially by a FLASH 2000 elemental analyzer (Thermo Fisher Scientific, MA, USA). Total phosphorus of plant and soil samples, and soil available phosphorus were used in molybdenum–antimony anti-spectrophotometric method. Soil available nitrogen was tested by Conway Method. Soil moisture measurement was in drying method, and soil pH and electrical conductivity were measured by PC2700 Desktop pH/Conductivity Measuring Instrument (Thermo Fisher Scientific, MA, USA). Leaf δ13C content was analyzed by Isoprime 100 (Elementar Analysensysteme, Germany). All analysis of samples was conducted in the laboratory of the China Agricultural University, Beijing.
One-way analysis of variance (ANOVA) was used to test significant differences in C%, N%, P%, C/N ratio, C/P ratio, N/P ratio and δ13C between degradation degrees or between plant species, respectively, and then followed by multiple comparison by Duncan’s post hoc test or the Games–Howell test for heterogeneous variances.
One-way analysis of variance (ANOVA) was used to test significant differences in C%, N%, P%, C/N ratio, C/P ratio, N/P ratio and δ13C between degradation degrees or between plant species, respectively, and then followed by multiple comparison by Duncan’s post hoc test or the Games–Howell test for heterogeneous variances.
3
DNA Carbon Isotope Analysis
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13C enrichment of DNA (total, heavy- and light-DNA fractions) was determined using an elemental analyzer coupled with an isotope ratio mass spectrometer (EA/IRMS) (VarioPyroCube and Isoprime 100, Elementar Analysensysteme GmbH, Langenselbold, Germany) as previously described by Haichar et al. (2007) [23 (link)]. 13C/12C ratios of DNA were expressed as δ in ‰ relative to V-PDB.
4
Stable Isotope Analysis of Dragonfly Wings
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We used a steam equilibration procedure to account for the exchangeable proportion of H and calculate the δ2H values of non-exchangeable H [37 (link), 38 (link)]. After the steam equilibration, stable hydrogen isotope ratios of dragonfly wings were measured with a TC/EA-IRMS device (vario PYRO Cube and IsoPrime 100, Elementar Analysesysteme GmbH, Germany). Further information on stable isotope analysis is provided in the supplementary information.
5
Carbon Isotope Analysis of Photosynthetic Leaves
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The leaves used for photosynthesis measurements (third and fourth from the top) were dried and ground in a ball mill (MM200, Retch, Germany). The triplicate of 2 mg samples was packed into a tin capsule for 13C isotope determination using a mass spectrometer (IRMS IsoPrime 100 Elementar, a vario PYRO cube, Elementar, Germany). The following formula was used to calculate δ13C (Kubásek et al., 2007 (link)):
where Rp is the 13C/12C obtained from a mass spectrometer in plant samples and Rs is a reference value of 13C/12C in standard V-PDB (Vienna Pee Dee Belemnite); all values were expressed as per mil of dry weight. Carbon isotope discrimination (Δ13C) in the leaves was calculated from plant δ13C values (δp) and air δ13C values (δa) with the following formula (Farquhar et al., 1989 ):
Bundle sheath leakiness (φ) was estimated according to Farquhar (1983) with the following expression:
where a, b3, b4, and s are isotopic discrimination constants; a (4.4‰) is CO2 in air diffusivity through the stomata, b3 (29‰) is the carboxylation of Rubisco, b4 (−5.7‰) is the HCO3– dissolution and fractionation of phosphoenolpyruvate (PEP) carboxylation, and s (1.8‰) is the leakage of CO2 from the bundle sheath to mesophyll cells.
where Rp is the 13C/12C obtained from a mass spectrometer in plant samples and Rs is a reference value of 13C/12C in standard V-PDB (Vienna Pee Dee Belemnite); all values were expressed as per mil of dry weight. Carbon isotope discrimination (Δ13C) in the leaves was calculated from plant δ13C values (δp) and air δ13C values (δa) with the following formula (Farquhar et al., 1989 ):
Bundle sheath leakiness (φ) was estimated according to Farquhar (1983) with the following expression:
where a, b3, b4, and s are isotopic discrimination constants; a (4.4‰) is CO2 in air diffusivity through the stomata, b3 (29‰) is the carboxylation of Rubisco, b4 (−5.7‰) is the HCO3– dissolution and fractionation of phosphoenolpyruvate (PEP) carboxylation, and s (1.8‰) is the leakage of CO2 from the bundle sheath to mesophyll cells.
6
Stable Isotope Analysis of Rice
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After weighing, the rice samples were ground into a fine powder and sieved (2mm), and 0.1 g of fine powdered rice organs was used for measurement of isotopic composition. Carbon concentration([C], %), carbon isotopic composition (δ13C, ‰) and nitrogen isotopic composition (δ15N, ‰) in plant organs were determined using an Elemental Analyzer System (vario PYRO cube, Elementar Analysensysteme GmbH, Germany) interfaced with an Isotope Mass Spectrometer (Isoprime 100, Elementar Analysensysteme GmbH, Germany). The carbon content in organ (Corgan) was calculated from the [C]organ and the dry biomass of the respective organ. The whole plant C content (Cwhole-plant) was calculated from the Cgrain, Cstem, Cleaf, and Croot. The C allocation (%) of organ was defined as the ratio of Corgan to Cwhole-plant. The δ13C and δ15N value of rice organs can be calculated as:
where R is the ratio of 13C/12C or 15N/14N.
The δ13C whole-plant was calculated as follows:
where R is the ratio of 13C/12C or 15N/14N.
The δ13C whole-plant was calculated as follows:
7
13CO2 Labeling of Crop Plants
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At the flowering stage, 10 plants from each genetic line/treatment were labeled with 13CO2. Flag leaves of the main stems were used for isotope labeling under ambient field conditions, which lasted for 30 min, between 9:00 a.m. and 11:00 a.m., on a clear day with photosynthetic active radiation at the top of the canopy ranging between 1000 and 1400 μmol·m−2·s−1. The whole flag leaf was individually enclosed in a polyethylene bag (25 cm length and 4 cm diameter) and exposed to a 13CO2-enriched atmosphere. 13C was supplied by the injection of 15 mL of 13CO2 gas (from Shanghai Research Institute of Chemical Industry Co., Ltd., Shanghai, China). At the maturity stage, the labeled plants were harvested, and each plant was divided into leaf blades, culms plus sheaths, and kernels. To determine total C and δ13C, samples were combusted in an elemental analyzer (CHNS elemental analyzer, Vario marco cube, Elementar, Germany) and an isotope ratio mass spectrometer (Isoprime100, Elementar, Germany). Isotopic values are expressed in d units, denoting parts per thousand deviations (‰) relative to VPDB (Vienna Pee Dee Belemnite).
8
Stable Isotope Analysis of Samples
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Approximately 2-2.5 mg of each sample was weighed and loaded into tin boats. All dried masses were measured to the nearest 0.01 mg. Stable isotope measurements were performed with an isotope ratio mass spectrometer (IsoPrime100) coupled to an N-C-S elemental analyser (Vario MICRO cube, Elementar) for automated analyses. Stable isotope abundances were expressed in delta (δ) notation as the deviation from standards in parts per thousand (‰) according to the following equation:
where X is 13 C, 15 N or 34 S and R is the corresponding ratio 13 C/ 12 C, 15 N/ 14 N or 34 S/ 32 S. The δ values are multiplied by 1000 for easier understanding. The isotopic ratios were estimated relative to the international standards of Vienna Pee Dee Belemnite (vPBD) for carbon, Atmospheric Air for nitrogen and Vienna Canyon Diablo Troilite (vCDT) for sulphur.
International Atomic Energy Agency certified reference materials (IAEA, Vienna, Austria)
were used as analytical standard calibrated against the international isotopic references sucrose (IAEA-C 6 , δ 13 C = -10.8 ± 0.5 ‰; mean ± SD), ammonium sulfate (IAEA-N 2 , δ 15 N = 20.3 ± 0.2 ‰; mean ± SD) and silver sulfide (IAEA-S 1 , mean δ 34 S = -0.3 ‰) as primary standards, and sulfanilic acid (δ 13 C= -25.6 ± 0.4 ‰; δ 15 N = -0.1 ± 0.5 ‰ ; δ 34 S= 5.9 ± 0.5 ‰ ; mean ± SD in each case) as secondary analytical standard.
where X is 13 C, 15 N or 34 S and R is the corresponding ratio 13 C/ 12 C, 15 N/ 14 N or 34 S/ 32 S. The δ values are multiplied by 1000 for easier understanding. The isotopic ratios were estimated relative to the international standards of Vienna Pee Dee Belemnite (vPBD) for carbon, Atmospheric Air for nitrogen and Vienna Canyon Diablo Troilite (vCDT) for sulphur.
International Atomic Energy Agency certified reference materials (IAEA, Vienna, Austria)
were used as analytical standard calibrated against the international isotopic references sucrose (IAEA-C 6 , δ 13 C = -10.8 ± 0.5 ‰; mean ± SD), ammonium sulfate (IAEA-N 2 , δ 15 N = 20.3 ± 0.2 ‰; mean ± SD) and silver sulfide (IAEA-S 1 , mean δ 34 S = -0.3 ‰) as primary standards, and sulfanilic acid (δ 13 C= -25.6 ± 0.4 ‰; δ 15 N = -0.1 ± 0.5 ‰ ; δ 34 S= 5.9 ± 0.5 ‰ ; mean ± SD in each case) as secondary analytical standard.
9
Leaf Nitrogen Content Determination
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Following the measurement of chlorophyll fluorescence parameters, two leaves from each plant were harvested, instantly frozen in liquid nitrogen, and preserved at -80°C for subsequent biochemical analyses. Another two leaves were heat-fixed at 105°C for 30 minutes and then dehydrated to constant mass at 65°C for the determination of leaf dry weight and leaf mass per unit area (LMA, g m-2) (n = 6).
Accurately weigh a certain amount of ground plant samples (2-3 mg) into tin foil cups, tightly wrap them, record the mass, and place them in the automatic sample introduction tray of the instrument for analysis using a stable isotope ratio mass spectrometer (Elementar, Isoprime 100, UK) to determine the N content (Nm, g/kg) in the leaves, stems, and roots. Leaf N content per unit leaf area (Narea, g m-2) was calculated by Nm × LMA (n = 6).
Accurately weigh a certain amount of ground plant samples (2-3 mg) into tin foil cups, tightly wrap them, record the mass, and place them in the automatic sample introduction tray of the instrument for analysis using a stable isotope ratio mass spectrometer (Elementar, Isoprime 100, UK) to determine the N content (Nm, g/kg) in the leaves, stems, and roots. Leaf N content per unit leaf area (Narea, g m-2) was calculated by Nm × LMA (n = 6).
10
Dark CO2 Fixation in Cocultures
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To verify dark CO2 fixation, 13C-labeled NaHCO3 was used as the source of CO2 in the coculture medium. When G. metallireducens and R. palustris cocultures were grown at their stationary phase, coculture cells were collected by centrifugation at 6000g for 10 min and then dehydrated using a vacuum freeze dryer (ALPHA 1-4 LDplus, Osterode, Germany). Dry cells (4 mg) were wrapped with tinfoil and placed into stable isotope ratio mass spectrometers (Isoprime 100, Elementar, Langenselbold, Germany) for dark 13CO2 fixation analysis following the manufacturers’ instructions.
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