2.2 Molecular methods to generate Angiosperms353 sequence data
Total DNA was isolated using a modified CTAB protocol (Doyle & Doyle,
1987). Genomic libraries were constructed as optimized in (Viruelet al. , 2019) using half volumes of the
NEBNext® UltraTM II DNA Library Prep Kit for
Illumina® (New England Biolabs, Ipswich, MA, United
States), purified using AMPure XP magnetic beads and multiplexed with
NEBNext® Multiplex Oligos for
Illumina® (Dual Index Primer Sets I and II). Equimolar
pools containing twelve genomic libraries were enriched with
half-reactions of the Angiosperms353 probe kit (Johnson et al. ,
2019; Baker et al. , 2022) following myBaits®kit manual v5.03 (Arbor Biosciences). DNA concentrations were calculated
using a QuantusTM fluorometer (Promega Corp.), and an Agilent 4200
TapeStation (Agilent Technologies, Santa Clara, CA, United States) was
used to assess fragment length. Sequencing was performed on a HiSeq
(Illumina, Inc.) by Macrogen (Seoul, South Korea), producing 150 bp
paired-end reads.