PacBio sequencing of the genome of Drosophila suzukii: adult female whole body
High-molecular weight DNA was extracted from 40 adult D. suzukiii females (WT3-2.0) using the Blood & Cell culture DNA midi kit (Qiagen). The quality and concentration of the DNA was assessed using a 0.5 % agarose gel (run for >8 h at 25 V) and a Nanodrop spectrophotometer (ThermoFisherScientific). PacBio libraries were generated using the SMRTbell Template Prep Kit 1.0 according to manufacturers instructions. In brief, 10 g of genomic DNA per library (estimated by Qubit assay) was sheared into 20 kb fragments using the Megaruptor system, followed by an exo VII treatment, DNA damage repair and end-repair before ligation of hair-pin adaptors to generate a SMRTbell library for circular consensus sequencing. The library was then subjected to exo treatment and PB AMPure bead wash procedures for clean-up before it was size selected with the BluePippin system (SAGE) with a cut-off value of 9000 bp. In total 48 units of SMRTcell with library was sequenced on the PacBio Sequel instrument using the Sequel 2.0 polymerase and 600 minute movie time.
SUB6662568
run5_F02