At_PEK_HiFi
SMRTbell target size libraries were constructed for sequencing according to PacBios standard protocol (Pacific Biosciences, CA, USA) using either 10 kb or 20 kb preparation solutions. The main steps for library preparation are: (1) gDNA shearing, (2) DNA damage repair, (3) blunt end-ligation with hairpin adapters from the SMRTbell Express Template Prep Kit 2.0 (Pacific Biosciences), (4) size selection, and (5) binding to polymerase. Briefly, a total amount of 2 g DNA per sample was used for the DNA library preparations. The genomic DNA sample was sheared by g-TUBEs (Covaris, USA) according to the expected size of the fragments for the library. Single-strand overhangs were then removed and DNA fragments were damage repaired, end polished, and ligated with the stem-loop adaptor for PacBio sequencing. Link-failed fragments were further removed by exonuclease and target fragments were screened by the BluePippin (Sage Science, USA). The SMRTbell library was then purified using AMPure PB, and Agilent 2100 Bioanalyzer (Agilent technologies, USA) was used to detect the size of library fragments.
At_PEK_HiFi