Preparation of environmental samples
For Field-Samples-1 data, white clover (Trifolium repens ) root
nodules were collected from two locations: Store Heddinge, Denmark (6
plots) and Aarhus University Science Park, Aarhus, Denmark (2 plots)
(Supplementary Figure S4 ). The clover varieties sampled were
Klondike (Store Heddinge) and wild white clover, (Aarhus). 100 large
pink nodules were collected from 4 points on each plot, making a total
of 32 samples. Nodules were stored at -20°C until DNA extraction. Nodule
samples were thawed at room temperature and crushed using a sterile
homogeniser stick. Crushed nodules were mixed with 750µl Bead Solution
from the DNeasy PowerLyzer PowerSoil DNA isolation kit (QIAGEN, USA) and
DNA was extracted following the manufacturer’s instructions. DNA sample
concentrations were measured using a Nanodrop 3300 instrument
(Thermofisher Scientific Inc., USA).
For Field-Samples-2 data, root nodules were additionally sampled from 13
white clover conventionally-managed field trial plots at Store Heddinge,
Denmark (Sample 1A-13A, Additional File 2 ). All plots were sown
under the same conditions in 2017. Three to ten clover plants were
sampled from one point in each plot and the 100 largest nodules
collected. Nodules were stored at -20°C, and DNA was extracted for each
sample using the Qiagen DNeasy PowerLyzer PowerSoil DNA isolation kit,
as above. Samples were processed independently with Platinum
(non-proofreading) and Phusion (proofreading) polymerases to evaluate
the method dependency on polymerase choice, as described in the
following sections.