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Development of an efficient Sanger sequencing-based assay for detecting SARS-CoV-2 spike mutations.
Lim, Ho Jae; Park, Min Young; Jung, Hye Soo; Kwon, Youngjin; Kim, Inhee; Kim, Dong Kwan; Yu, Nae; Sung, Nackmoon; Lee, Sun-Hwa; Park, Jung Eun; Yang, Yong-Jin.
  • Lim HJ; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Park MY; Department of Integrative Biological Sciences & BK21 FOUR Educational Research Group for Age-associated Disorder Control Technology, Chosun University, Gwangju, Republic of Korea.
  • Jung HS; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Kwon Y; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Kim I; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Kim DK; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Yu N; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Sung N; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Lee SH; Clinical Research Institute, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Park JE; Department of Molecular Diagnostics, Seegene Medical Foundation, Seoul, Republic of Korea.
  • Yang YJ; Department of Integrative Biological Sciences & BK21 FOUR Educational Research Group for Age-associated Disorder Control Technology, Chosun University, Gwangju, Republic of Korea.
PLoS One ; 16(12): e0260850, 2021.
Article in English | MEDLINE | ID: covidwho-1613341
ABSTRACT
Novel strains of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) harboring nucleotide changes (mutations) in the spike gene have emerged and are spreading rapidly. These mutations are associated with SARS-CoV-2 transmissibility, virulence, or resistance to some neutralizing antibodies. Thus, the accurate detection of spike mutants is crucial for controlling SARS-CoV-2 transmission and identifying neutralizing antibody-resistance caused by amino acid changes in the receptor-binding domain. Here, we developed five SARS-CoV-2 spike gene primer pairs (5-SSG primer assay; 69S, 144S, 417S, 484S, and 570S) and verified their ability to detect nine key spike mutations (ΔH69/V70, T95I, G142D, ΔY144, K417T/N, L452R, E484K/Q, N501Y, and H655Y) using a Sanger sequencing-based assay. The 5-SSG primer assay showed 100% specificity and a conservative limit of detection with a median tissue culture infective dose (TCID50) values of 1.4 × 102 TCID50/mL. The accuracy of the 5-SSG primer assay was confirmed by next generation sequencing. The results of these two approaches showed 100% consistency. Taken together, the ability of the 5-SSG primer assay to accurately detect key SARS-CoV-2 spike mutants is reliable. Thus, it is a useful tool for detecting SARS-CoV-2 spike gene mutants in a clinical setting, thereby helping to improve the management of patients with COVID-19.
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Full text: Available Collection: International databases Database: MEDLINE Main subject: Sequence Analysis, RNA / Spike Glycoprotein, Coronavirus / SARS-CoV-2 / Mutation Type of study: Diagnostic study / Prognostic study Limits: Humans Language: English Journal: PLoS One Journal subject: Science / Medicine Year: 2021 Document Type: Article

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Sequence Analysis, RNA / Spike Glycoprotein, Coronavirus / SARS-CoV-2 / Mutation Type of study: Diagnostic study / Prognostic study Limits: Humans Language: English Journal: PLoS One Journal subject: Science / Medicine Year: 2021 Document Type: Article