Sorted population for BSA: CTS1 tagged
Library preparation for Illumina sequencing was performed using Nextera DNA Library Prep kit (Illumina) with modifications. The tagmentation was done on 5 ng of DNA using 4 l of Tagment DNA buffer (TD in the kit) and 0.25 l of Tagment DNA enzyme (corresponding to a 20-fold dilution of TDE1 from the kit) and incubating for 10 minutes at 55C. Fragments were amplified with index primers (8 Nextera primers i5 and 12 Nextera primers i7, for up to 96 possible multiplex combinations) on 10 l tagmented DNA by adding 1 l of each primer solution (10 M), 5 l of 10X ExTaq buffer and 0.375 l of ExTaq polymerase (Takara) and water to a final volume of 50 l. The amplification was run for 17 PCR cycles (95C denaturation, 62C annealing, 72C elongation). 10 l of each reaction were pooled for multiplexing and run on a 2% agarose gel. DNA that migrated between the 400 and 600 bp was extracted using Monarch DNA Gel Extraction Kit (NEB T1020L). The pooled library DNA concentration was determined using Qubit dsDNA BR Assay Kit (ThermoFisher Scientist Q32853), and submitted for sequencing. Sequencing was performed at the University of Minnesota Genomics Core (UMGC). Our 125 populations were processed in four batches extracted and sequenced at different times. Two were sequenced using an Illumina HiSeq 2500 (high-output mode; 50-bp paired-end) and two were sequenced using an Illumina NextSeq 500 (mid-output mode, 75-bp paired-end).