2.2. Whole-genome sequencing of SARS-CoV-2
We conducted whole-genome sequencing (WGS) of SARS-CoV-2 for
RT-PCR-positive samples with a Ct value of ≤30 based on our previous
studies.30-33,36 Briefly, SuperScript™ III
First-Strand Synthesis System (Thermo Fisher Scientific, MA, United
States) was used to synthesize single-stranded complementary DNA (cDNA)
from the total RNA. Subsequently, the second strand was synthesized
using Covid-19 ARTIC v3 primer pool design by SARS-CoV-2 ARTIC Network
using Phusion™ High-Fidelity DNA Polymerase (Thermo Fisher Scientific,
MA, United States). The library preparations were performed using the
Illumina DNA Prep (Illumina, California, United States). The Illumina
MiSeq next-generation sequencer was used to perform the whole-genome
sequencing of SARS-CoV-2. The genomes of our samples were assembled and
mapped into the reference genome from the first Wuhan (China) isolate
(hCoV-19/Wuhan/Hu-1/2019, GenBank accession number: NC_045512.2) using
Burrow-Wheeler Aligner (BWA) algorithm embedded in UGENE v. 1.30
(https://doc.ugene.net/wiki/display/UUOUM30/About+UGENE). The
lineage of each sequence was evaluated using Pangolin v4.1.2. The GISAID
ID of each submitted isolate generated from our study was shown in
Supplementary Table 1.