Next-generation sequencing
Next-generation sequencing (NGS) was conducted using Illumina MiSeq for every round of DNA library (0 to 4 rounds). Prior to sequence library preparation, quantification was carried out using Qubit 2.0 Fluorometer and Qubit 1X dsDNA HS Assay Kit (Thermo Fisher Scientific). A total 50 ng of the DNA library sample was used as an input for the NEBNext Ultra II DNA Library Prep with Sample Purification Beads (NEB). We also used NEBNext Multiplex Oligos for Illumina (Index Primers Set 1) (NEB) to multiplex our samples. The length of prepped DNA Library was determined by using Agilent 2100 Bioanalyzer (Agilent Technologies), quantified using Qubit 2.0 Fluorometer and run on MiSeq illumina platform using the Miseq Reagent Kit v3 600 cycles.