PacBio RS II sequencing
N/A
Diploid cells of the bioethanol isolate Saccharomyces cerevisiae PE-2 (Basso et al. FEMS Yeast Res. 2008 Nov;8(7):1155-63.) were induced to sporulate. A single tetrad was dissected and two spores were isolated. Those correspond to the spore H3 (MATα) and H4 (MATa). Individual cell cultures were prepared from each spore and high quality DNA was extracted (DNeasy Plant Mini Kit, QIAGEN, Germany) from each sample. The isolated DNA was shipped to the DNA link company (Seoul, Republic of Korea) for library construction and sequencing. A total of 5μg for each sample was used as input into library preparation. The SMRTbell library was constructed with SMRTbell™ Template Prep Kit 1.0 (PN 100-259-100) following manufacture's instructions (Pacific Biosciences) with selection for fragments higher than 10 kb. SMRTbell library was sequenced using 1 SMRT cells (Pacific Biosciences) using C4 chemistry (DNA sequencing Reagent 4.0) and 240 minute movies were captured for each SMRT cell using the PacBio RS II (Pacific Biosciences) sequencing platform. The resulting reads were filtered for quality (≥ 0.75) and length (≥ 50 nts) according to default filtering parameters defined by Pacific Biosciences.