Construction of DNA-NanoLuc luciferase conjugates for DNA aptamer-based sandwich assay using Rep protein
| Autor: | Masayasu Mie, Takahiro Niimi, Yasumasa Mashimo, Eiry Kobatake |
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| Rok vydání: | 2019 |
| Předmět: |
Circovirus
0106 biological sciences 0301 basic medicine Aptamer Gene Expression Bioengineering 01 natural sciences Applied Microbiology and Biotechnology Viral Proteins 03 medical and health sciences chemistry.chemical_compound 010608 biotechnology Escherichia coli A-DNA Luciferase Luciferases Luminescent Agents Staining and Labeling biology Chemistry DNA Helicases General Medicine Aptamers Nucleotide biology.organism_classification Porcine circovirus 030104 developmental biology Biochemistry Trans-Activators Replication initiator protein DNA Biotechnology Conjugate |
| Zdroj: | Biotechnology Letters. 41:357-362 |
| ISSN: | 1573-6776 0141-5492 |
| DOI: | 10.1007/s10529-018-02641-7 |
| Popis: | We developed a DNA-NanoLuc luciferase (NnaoLuc) conjugates for DNA aptamer-based sandwich assay using the catalytic domain of the replication initiator protein derived from porcine circovirus type 2 (pRep). For construction of DNA aptamer and NanoLuc conjugate using the catalytic domain of Rep from PCV2. pRep fused to NanoLuc was genetically constructed and expressed in E. coli. After purification, the activities of fused pRep and NanoLuc were evaluated, and DNA-NanoLuc conjugates were constructed via the fused pRep. Finally, constructed DNA-NanoLuc conjugates were applied for use in a DNA aptamer-based sandwich assay. Here, pRep was used not only for conjugation of the NanoLuc to the detection aptamer, but also for immobilization of the capture aptamer on the plate surface. We have demonstrated that DNA-NanoLuc conjugates via the catalytic domain of PCV2 Rep could be applied for DNA aptamer-based sandwich assay system. |
| Databáze: | OpenAIRE |
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