Element AVITI Long Insert  Sequencing of the GIAB HG008-N-D normal duodenal tissue
Sequencing was performed on the HG008-T pancreatic ductal adenocarcinoma tumor cell line with matched normal duodenal tissue (HG008-N-D) and normal pancreatic tissue (HG008-N-P), using similar methods and library preparation protocols tailored for both tumor and normal samples as described below. The data presented here corresponds to the normal duedonal 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.Library preparation was performed using  isolated genomic DNA (gDNA) from non-viable HG008-T cells (batch 0823p23) and non-viable HG008-N-D and HG008-N-P tissue.  One long insert library (>1kb) was prepared from each gDNA source.  For each library, one microgram of gDNA was input into the Element Elevate Mechanical Library Prep Kit (PN: 830-00008, Document # MA-00004 Rev. E) using the Elevate Long UDI Adapter Kit Set A (PN: 830-00010).  Mechanical shearing of gDNA was performed using a Covaris ultra-sonicator to achieve a longer insert size of ~800-900 bp. For the long insert libraries one iteration with the following settings were performed: 5 second duration, peak power 50, duty cycle 20, 1000 cycles per burst, average power 10. PCR-free library preparation continued with end-repair, A-tailing and adapter ligation, generating dual-index libraries. Size selection was performed with double-sided SPRI bead clean-up with SPRI ratios of 0.3X/0.42X.  Linear libraries were quantified for loading onto the AVITI system using qPCR using the 2x Phanta Flash master mix (Vazyme PN: P510) and Evagreen (Biotium PN:31000).