Truseq cd indexes

Total genomic DNA was extracted from fresh leaves of ‘Akiou', ‘Fuyu', and ‘Taishuu' using the cetyl trimethyl ammonium bromide method and purified by phenol/chloroform extraction. Then, 1.5 μg of DNA from each sample was used to prepare short-read genomic libraries according to the following steps. First, DNA molecules were enzymatically fragmented and size-selected to 400–800 bp using AMPure (0.4:1 v/v AMPure: reaction). The resulting fragments were subjected to genomic library construction using the GenNext NGS Library Prep kit (Toyobo, Japan) and Truseq CD indexes (Illumina, USA) according to the manufacturers’ instructions and pooling guide. After amplification, libraries were further refined to remove fragments longer than 800 bp and shorter than 300 bp by AMPure. Finally, three genomic sequence libraries, with a gradient insert size of 300–800 bp, were mixed in equal proportions and sequenced using two lanes of the Illumina HiSeqX PE 150 platform (Illumina, San Diego, CA, USA). A total of 127.67, 93.47, and 68.73 Gb of raw DNA sequencing data (approximately 20-fold coverage of the polyploid genomes) of ‘Akiou', ‘Fuyu', and ‘Taishuu', respectively, were used for further analysis.