Abstract
The novel SARS-CoV-2 outbreak has swiftly spread worldwide. The rapid genome sequencing of SARS-CoV-2 strains has become a helpful tool for better understanding the genomic characteristics and origin of the virus. To obtain virus whole-genome sequences directly from clinical specimens, we performed nanopore sequencing using a modified ARTIC protocol in a portable nanopore sequencer and validated a routine 8-h workflow and a 5-h rapid pipeline. We conducted some optimization to improve the genome sequencing workflow. The sensitivity of the workflow was also tested by serially diluting RNA from clinical samples. The optimized pipeline was finally applied to obtain the whole genomes of 29 clinical specimens collected in Hangzhou from January to March 2020. In the 29 obtained complete genomes of SARS-CoV-2, 33 variations were identified and analyzed. The genomic variations and phylogenetic analysis hinted at multiple sources and different transmission patterns during the COVID-19 epidemic in Hangzhou, China. In conclusion, the genomic characteristics and origin of the virus can be quickly determined by nanopore sequencing following our workflows.
Highlights
The novel SARS-CoV-2 outbreak has swiftly spread worldwide
To acquire the whole-genome sequence of SARS-CoV-2 more efficiently, an 8-h workflow was designed on the basis of the sequencing throughput and speed after loading the library into the flow cell, and a 5-h workflow was designed for rapid library building (Fig. 1)
These two workflows were both tested on the HZCDC0001 sample, with Ct values of 26.51/27.03 (Orf1ab/N); this sample was obtained from the first case that appeared in Hangzhou, Zhejiang Province
Summary
The rapid genome sequencing of SARS-CoV-2 strains has become a helpful tool for better understanding the genomic characteristics and origin of the virus. To obtain virus whole-genome sequences directly from clinical specimens, we performed nanopore sequencing using a modified ARTIC protocol in a portable nanopore sequencer and validated a routine 8-h workflow and a 5-h rapid pipeline. The optimized pipeline was applied to obtain the whole genomes of 29 clinical specimens collected in Hangzhou from January to March 2020. We applied a modified ARTIC protocol for SARS-CoV-2 genome sequencing on the MinION platform. Two workflows were applied and validated by amplifying and sequencing the genomes from the clinical samples of SARS-CoV-2-infected patients, and the characteristics of 29 SARS-CoV-2 genomes collected in Hangzhou were analyzed to study the origin and transmission history of these viruses
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