Sampling and analyses
Considering the effect of fast N transformation (Holst et al. 2007; Song
et al. 2007), plant community sampling and species sampling were carried
out in each experimental plot 24 hours after injection. A 0.5 × 0.5 m
quadrat was selected, and the aboveground part of the plant was sampled
by the harvesting method. Root and soil were sampled and merged using a
root drill (5 cm in diameter), ”Z”-shaped, 0–30 cm deep, with five
drills. This depth contains 95% of the roots in the alpine meadow. The
infiltration depth of the injection solution was not more than 10 cm
based on a preliminary test. The root samples were separated from the
soil and washed with water, and the soil samples were sifted through a
sieve (<2 mm) and then air-dried.
Species sampling was carried out in the remaining plot areas after
community sampling, and plants that were common to each quadrat were
sampled. Seven species were common to each quadrat in the undegraded
alpine meadow, including E. nutans , P. annua , K.
humilis , S. distigmaticus, P. anserina, P. kansuensis,and L. brachystachya . There were four common species in each
degraded alpine meadow quadrat, including E. nutans , P.
annua , P. kansuensis, and Ajania tenuifolia . The other
plant species were not collected by species owing to their uneven
distribution, though they were dominant species in the degraded alpine
meadow. Among them, E. nutans , P. annua , and P.
kansuensis were common generalist species in the two alpine meadows,
and the others were unique species in the two meadow types. We collected
6–12 plants from each species and separated the shoots from the roots.
In the laboratory, all plant samples were washed with distilled water to
remove 15N residue on the surface and dried for 48
hours in a 70°C oven. The powder was then ground, and the15N and N concentrations were analyzed by isotope mass
spectrometry. The soil 15N and total N concentrations
were analyzed by isotope mass spectrometry, the concentrations of
NO3- and
NH4+ were determined using an
autoanalyzer (AA3, Branl- Luebbe, Germany) in 0.5 M
K2SO4 extracts, and soluble organic N
was determined according to the method described by Chen et al. (2017).