Experimental set-up
From a pre-experiment, we known that species germination rates were
different. Therefore, we sowed them on different date (Table S1) to
ensure that all species had a similar developmental stage at the start
of the experiment. From 26 June to 14 August 2020 (Table S1), we used
plastic trays (19.5 cm × 14.6 cm × 6.5 cm) which filled with peat moss
as subtrate (Pindstrup Plus, Pindstrup Mosebrug A/S, Denmark) to sow
seeds and transplanted stolons of each plant, and placed all trays in a
greenhouse under natural light conditions, with a temperature between 20
°C and 28 °C.
On 21 September 2020, we collected the field soil from different sites,
a natural grassland site (Changling: 123°30’32.26”E, 44°33’21.74”N) and
two abandoned farmland sites (Anhui: 117°12’45.58”E, 31°55’7.68”N;
Taizhou: 121°40’43.60”E, 19°1’0.12”N), where the test plants occur
commonly. We collected soil at a depth of 0-25 cm in each site, and then
used a 0.5 cm mesh to sieved and homogenized the three sites at a ratio
of 1:1:1. Then we stored half of the soil at 4 °C until pot-filling. The
other half of the soil we sterilized four days with a dose of 25 kGy of60COγ irradiation at the Harbin
Guangya Radiation New Technology Co., Ltd (Harbin, China). To ensure
that plants can fully grow, we homogenized 5 g slow-release fertilizer
(Osmocote® Exact Standard, Everris International B.V., Geldermalsen, The
Netherlands) with the substrate in each pot.
We used 2.5 L circular pots (top diameter × bottom diameter × height:
18.5 cm × 12.5 cm × 15 cm) and filled with a mixture of 37.5% (v/v)
sand, 37.5% (v/v) vermiculite and 25% live or sterilized field soil.
On 29 August 2020, we selected the similar seedlings for each species,
and transplanted at total of 20 seedlings into 20 pots (i.e., one
seedling per pot, see Figure 1). The pot size was selected for that the
size was large enough to avoid that growth restriction until the end of
the experiment. To test the effects of soil microbes on invasive and
native plant species at different time periods, we assigned the 20 pots
of each species to two levels of soil (live vs sterilized)
treatments fully crossed with two times of harvest
(first vs second). In other
words, per species, we had five pots (i.e., replicates) in each of the
four treatment combinations, 160 pots in total (2 levels of plant
invasion status [invasive vs native] × 4 plant species pairs
× 2 soil [live vs sterilized] × 2 harvest time
(first vs second) × 5
replicates). The resulting 160 pots were randomly assigned to the four
shelves in the greenhouse under natural light conditions, with a
temperature between 20 °C and 28 °C, and rerandomized again after six
weeks (i.e., on 8 October 2020). We gave each pot a separate tray in
order to avoided the loss of nutrients and water, and also avoided soil
biota cross contamination. We checked the pots every day and watered
fully. It was noted that 80 pots of the first harvest were shared for
data with another experiment (reported in Zhang et al. 2022).
We had twice harvest on 10 November and 2 December 2020, respectively.
For each harvest, we separately harvested the above-ground biomass and
below-ground biomass of all plants, as one plant transplanted error and
four plants experienced drought, the total number of harvested plants
was 155. All above-ground biomass and below-ground biomass were dried
for at least 72 h at 65 °C, and then weighed. We calculated total
biomass (i.e., above-ground biomass + below-ground biomass) and root
mass fraction (RMF; below-ground biomass/total biomass).