WGS of Heterodera glycines MM26B
Genomic DNA was extracted using a Zymo Quick-DNA HMW MagBead Kit (Cat# D6060; Zymo Research Corp, Irvine, CA) following the manufacturers instructions. On average, each pool yielded 11 to 14 ng of DNA in 50 L of elution buffer (Zymo Research Corp). DNA quality was estimated for each pool and met the required high quality purity criteria of A260/A280 ~1.8 using a BioSpectrometer (Eppendorf, Hamburg, Germany). Prior to library preparation, each sample was normalized to 1.8 ng/L of DNA; and 25 L of each normalized DNA sample was used for library construction. Briefly, paired-end (PE) libraries were prepared using a KAPA HyperPrep Kit for DNA Library Preparation (Cat# KK8504; Kapa Biosystems, Wilmington, MA) following the kits protocol. The eight libraries were bead-based cleansed before their quality was checked for library concentration as well as size distribution, using a Qubit Fluorometer (Invitrogen, Carlsbad, CA), qPCR with a KAPA Library Quantification Kit (Cat# KK4854; Kapa Biosystems), and a Fragment Analyzer (Agilent Technologies, Santa Clara, CA). Libraries were pooled equimolarly to 5 nM in a final pool, and quality-controlled again with qPCR, Qubit reading, and fragment analysis prior to loading to the sequencer. Pool sequencing was performed on an Illumina NextSeq 2000 platform using the P3 PE 150 protocol (i.e., the data read length is pair-end-150 length). To meet our sequencing goal to reach a 150X coverage at the pooled (population) level, a single lane (flowcell) was used and yielded approximately 189X coverage. Library preparation and Pool sequencing were conducted by the Georgia Genomics and Bioinformatics Core at the University of Georgia.