2.4 Soil sampling and measurements
Soil samples were collected 30 times throughout the experimental period,
especially focusing on the crop growing season approximately every 2
weeks. For each sample, five composite soil samples (0–15 cm depth)
were taken inside the plot (7 m × 4 m; avoiding the plot edge, and c.a.
1 m away from the CO2 chambers mentioned below), and
between plants (plant-to-plant distance was 30 cm) so as not to disturb
plant roots. After transporting to the laboratory in a 4 °C cooler, soil
samples were passed through a 4-mm sieve after removing stones and plant
roots and stored at 4°C under field-moisture conditions until each
measurement. SOC was measured at the start of the experiment (in Sep
2017) and at the end of the experiment (in Dec 2019). Soil moisture
content was determined by the difference in soil weight before and 48 h
after 105 °C drying. MBC was measured using the fumigation-extraction
method (Vance et al., 1987), following the detailed described in
Sugihara, et al., 2015. All data were expressed on a dry weight basis.
To determine the soil bulk density, soil cores were also collected at
the start of cultivation and at the end of cultivation every year, i.e.,
in Sep 2017, Jan 2018, Aug 2018, Dec 2018, Aug 2019, and Dec 2019, only
in the C and B treatments. Five core samples were collected for each
sample by inserting metal rings of 100 cm3.