title: Long read-Seq of synthetic S. cerevisiae strains constructed during the project "Building synthetic chromosomes from natural DNA"
design: For ONT library preparation, the SQK-LSK109 and EXP-NBD104 protocols were used with the following modifications. At the end of each bead clean up, DNA was eluted in EB at 37oC for 10 mins to allow for the recovery of larger molecules. The library was then barcoded and multiplexed with up to 10 samples. These multiplexes were sequenced on R9.4.1 Chemistry, MIN-106 flow cells. The .fast5 archive was Tarbelled, gunzipped, and uploaded onto the USCs Center for Advanced Research Computing (CARC) Discovery cluster. Basecalling was performed using Guppy (v. 6.0.1), which was contained in a Singularity container using the configuration dna_r9.4.1_450bps_sup.cfg with 16 threads and a V100 GPU. The .fastq files were demultiplexed and trimmed of barcodes in Guppy using standard parameters.