| Autor: |
Francesca Marino-Merlo, Valeria Stefanizzi, Agnese Ragno, Lucia Piredda, Sandro Grelli, Beatrice Macchi, Antonio Mastino |
| Jazyk: |
angličtina |
| Rok vydání: |
2022 |
| Předmět: |
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| Zdroj: |
Life, Vol 12, Iss 8, p 1130 (2022) |
| Druh dokumentu: |
article |
| ISSN: |
2075-1729 |
| DOI: |
10.3390/life12081130 |
| Popis: |
Based on previous experience in our laboratory, we developed a real-time reverse transcriptase (RT) quantitative PCR (RT-qPCR) assay for the assessment of very low levels of HIV-1 RT activity. The RNA, acting as a template for reverse transcription into cDNA by HIV-1 RT, consisted of a synthetic RNA ad hoc generated by in vitro transcription and included a coding sequence for HSV-1 gD (gD-RNA-synt). Different conditions of variables involved in the RT-qPCR reaction, notably different amounts of gD-RNA-synt, different mixes of the reaction buffer, and different dNTP concentrations, were tested to optimize the assay. The results indicated that the gD-RNA-synt-based RT assay, in its optimized formulation, could detect a specific cDNA reverse transcription even in the presence of 1 × 10−9 U of HIV RT. This achievement greatly improved the sensitivity of the assay over previous versions. In summary, this constructed RT-qPCR assay may be considered a promising tool for providing accurate information on very low HIV-1 RT activity. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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