title: Novel system for detecting SARS coronavirus nucleocapsid protein using an ssDNA aptamer.
abstract: The outbreak of severe acute respiratory syndrome (SARS) in 2002 affected thousands of people and an efficient diagnostic system is needed for accurate detection of SARS coronavirus (SARS CoV) to prevent or limit future outbreaks. Of the several SARS CoV structural proteins, the nucleocapsid protein has been shown to be a good diagnostic marker. In this study, an ssDNA aptamer that specifically binds to SARS CoV nucleocapsid protein was isolated from a DNA library containing 45-nuceotide random sequences in the middle of an 88mer single-stranded DNA. After twelve cycles of systematic evolution of ligands by exponential enrichment (SELEX) procedure, 15 ssDNA aptamers were identified. Enzyme-linked immunosorbent assay (ELISA) analysis was then used to identify the aptamer with the highest binding affinity to the SARS CoV nucleocapsid protein. Using this approach, an ssDNA aptamer that binds to the nucleocapsid protein with a K(d) of 4.93±0.30nM was identified. Western blot analysis further demonstrated that this ssDNA aptamer could be used to efficiently detect the SARS CoV nucleocapsid protein when compared with a nucleocapsid antibody. Therefore, we believe that the selected ssDNA aptamer may be a good alternative detection probe for the rapid and sensitive detection of SARS.
authors.affiliation: Department of Bio and Nanochemistry, Kookmin University, Seoul 136-702, Republic of Korea.
authors.forename: Seong-Je
authors.initials: SJ
authors.lastname: Cho
authors.forename: Hye-Min
authors.initials: HM
authors.lastname: Woo
authors.forename: Ki-Sun
authors.initials: KS
authors.lastname: Kim
authors.forename: Jong-Won
authors.initials: JW
authors.lastname: Oh
authors.forename: Yong-Joo
authors.initials: YJ
authors.lastname: Jeong
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000378:metabolism; D001483:Base Sequence; D015153:Blotting, Western; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins / Q000235:genetics / Q000302:isolation & purification*; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000302:isolation & purification*; D052156:SELEX Aptamer Technique / Q000379:methods*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D045169:Severe Acute Respiratory Syndrome / Q000175:diagnosis
chemical_list: D052157:Aptamers, Nucleotide; D004277:DNA, Single-Stranded; D019590:Nucleocapsid Proteins