Huizao_Pacbio (CRX1331872; CRA021913)
To prepare DNA for the long-read sequencing, high molecular weight genomic DNA was extracted using the SDS technique and purified using the QIAGEN Genomic kit (Cat#13343, QIAGEN). On 1% agarose gels, the level of DNA degradation and contamination was evaluated. Using a NanoDrop One UV-Vis spectrophotometer (Thermo Fisher Scientific, USA), the OD 260/280 and 260/230 ranges were determined to be between 1.8 and 2.0 and 2.0 and 2.2, respectively. Pacbio HiFi SMRTbell libraries were constructed following the standard protocol using the SMRTbell Express Template Prep Kit 2.0 (PacBio, CA, USA). The lengthy DNA fragments were skillfully sheared down to 15-18 kb using a g-TUBE (Covaris, MA, USA). Single-strand overhangs were cut off and damaged and broken DNA was patched up with the chemicals in the Template Prep Kit. Once the ends were fixed, SMRTbell hairpin adapters were ligated to them, and then the libraries were concentrated and purified using AMPure PB beads (PacBio, CA, USA). BluePippin was utilized to size-select SMRTbell templates more than 15 kb to get large-insert SMRTbell libraries for sequencing (SageScience, MA, USA). The sequencing was performed using a PacBio Sequel II device with Sequencing Primer V2 and a Sequel II Binding Kit 2.0. For the raw sequencing reads, the min passes = 3 and min RQ = 0.99 default parameters in CCS software were utilized to generate high-precision HiFi reads with quality over Q20. submitted_instrument_model: PacBio Sequel