HIFI-Seq of Phthonandria atrilineata: adult mussle
High-molecular-weight genomic DNA was extracted from adult Phthonandria atrilineata specimens using a modified CTAB method. The quality and integrity of the extracted DNA were assessed using pulsed-field gel electrophoresis (PFGE) and Qubit fluorometry to ensure high molecular weight (>50 kb) and sufficient quantity for HiFi sequencing.The SMRTbell library was prepared following PacBio's protocol for the SMRTbell Express Template Prep Kit 2.0. Briefly, the genomic DNA was first sheared to an average size of 15-20 kb using g-TUBE (Covaris). The sheared DNA underwent damage repair and end-repair processes to create blunt ends. Afterwards, overhang adapter ligation was performed to create SMRTbell templates.The library was size-selected using the BluePippin system (Sage Science) with a 15 kb cut-off to enrich for longer fragments, which is crucial for generating HiFi reads. The size-selected library was then subjected to additional damage repair to ensure high library quality.Library quality was assessed using a Femto Pulse system (Agilent) to confirm the size distribution and quantity. The final library concentration was determined using a Qubit fluorometer.Sequencing was performed on a PacBio Sequel II system using Sequel II Binding Kit 2.0 and Sequel II Sequencing Kit 2.0. The library was bound to the sequencing polymerase using a 10-hour movie time. The sequencing run was set up to generate HiFi reads, which involves multiple passes of the same circular template to produce high-accuracy long reads.
SUB14765118
PHAT_HIFI_2024