Virology
Effects of sample processing modes on high-throughput sequencing of coronavirus whole genome
Chen Jing, Zhang Yawei, Niu Peihua, Lu Rojian, Zhu Na, Fan Hang, Tan Wenjie
Published 2020-02-29
Cite as Chin J Microbiol Immunol, 2020,40(02): 103-109. DOI: 10.3760/cma.j.cn112309-20191011-00322
Abstract
ObjectiveTo study the effects of different pre-sequencing sample processing modes on the results of whole genome sequencing with high-throughput sequencing (HTS) by taking the largest RNA virus (human coronavirus, HCoV) as the representative.
MethodsCell-cultured human coronavirus HCoV-OC43 strains were used as the representative samples and divided into different groups based on pre-sequencing processing modes as follows: untreated group, DNase and RNase treatment before nucleic acid extraction group, DNase treatment after nucleic acid extraction group, and DNase and RNase treatment before nucleic acid extraction and DNase treatment after nucleic acid extraction group. Nucleic acid samples of each group were analyzed by direct RNA sequencing (without amplification) and DNA sequencing after sequence independent single primer amplification (SISPA), respectively.
ResultsNo significant difference in viral genome coverage rates was observed between different groups. The highest genome coverage and sequencing accuracy were obtained in DNase treatment after nucleic acid extraction group by direct RNA sequencing, and the ratio of viral reads and the sequencing depth of each locus were effectively improved by SISPA amplification.
ConclusionsThis study provided an optimized technical strategy for whole genome sequencing of RNA viruses such as coronavirus.
Key words:
Human coronavirus; Sequence independent single primer amplification; Whole genome sequencing; High-throughput sequencing
Contributor Information
Chen Jing
Institute of Medical Virology, Zhejiang Provincial Key Laboratory of Medical Genetics, Key Laboratory of Laboratory Medicine of Ministry of Education, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China
Zhang Yawei
State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, China
Niu Peihua
NHC Key Laboratory of Biosafety, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
Lu Rojian
NHC Key Laboratory of Biosafety, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
Zhu Na
NHC Key Laboratory of Biosafety, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
Fan Hang
State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, China
Tan Wenjie
Institute of Medical Virology, Zhejiang Provincial Key Laboratory of Medical Genetics, Key Laboratory of Laboratory Medicine of Ministry of Education, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China
