title: Ultima UG100 ppmSeq Sequencing of the GIAB HG008-N-D duodenal tissue
design: Sequencing was performed on the HG008-T pancreatic ductal adenocarcinoma tumor cell line with matched normal duodenal tissue (HG008-N-D) using similar methods and library preparation protocols as described below. The data presented here corresponds to the normal duodenal tissue (HG008-N-D). Genomic DNA (gDNA) from non-viable normal tissue was isolated using the Qiagen Puregene Tissue Kit (PN: 158667).  Approximately 2 ml of Qiagen kit cell lysis buffer with 30 ul of Proteinase K was added to 10-15 mg of tissue. A Dounce Homogenizer was used to disrupt the tissue before leaving it overnight at 55C. This was followed by addition of RNase A and another incubation at 37C for 5 minutes followed by cooling on ice. Genomic DNA was precipitated, washed, and hydrated in TE buffer (1 mM EDTA, 10 mM TrisCl pH 7.5) per the Qiagen Puregene protocol. One paired plus minus sequencing (ppmSeq) library was prepared starting with 250 ng of isolated HG008-N-D gDNA. PpmSeq library preparation was performed using the NEBNext Ultra II DNA PCR-free Library Prep Kit (PN: E7410) following the Ultima ppmSeq protocol (UG ppmSeq Genomic DNA Library preparation Protocol for Solaris Free D100093 Rev.02), UG ppmSeq Adapters (96-sample indices plate) (PN: UG0105) for UG 100 Sequencer compatibility were used. Sequencing was performed on the UG 100 Sequencer (software version : APL 5.1.0.19) using iV27 chemistry. A human genome control sample HG002 (Coriell Nucleic acid ID: NA24385) was spiked into each run in order to ensure quality of the samples being sequenced. Libraries were pooled and single-end, approximately 200 bp reads were generated for both samples using the UG_116cycles_Baseline_1.7.0.0 sequencing recipe and flow order [T,G,C,A] on a single wafer.