A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock.
One of the biggest challenges during the pandemic has been obtaining and maintaining critical material to conduct the increasing demand for molecular tests. Sometimes, the lack of suppliers and the global shortage of these reagents, a consequence of the high demand, make it difficult to detect and d...
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2021-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0248885 |
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| author | Adolfo Marcelo Ñique Fiorella Coronado-Marquina Jairo Andrés Mendez Rico María Paquita García Mendoza Nancy Rojas-Serrano Paulo Vitor Marques Simas Cesar Cabezas Sanchez Jan Felix Drexler |
| author_facet | Adolfo Marcelo Ñique Fiorella Coronado-Marquina Jairo Andrés Mendez Rico María Paquita García Mendoza Nancy Rojas-Serrano Paulo Vitor Marques Simas Cesar Cabezas Sanchez Jan Felix Drexler |
| author_sort | Adolfo Marcelo Ñique |
| collection | DOAJ |
| description | One of the biggest challenges during the pandemic has been obtaining and maintaining critical material to conduct the increasing demand for molecular tests. Sometimes, the lack of suppliers and the global shortage of these reagents, a consequence of the high demand, make it difficult to detect and diagnose patients with suspected SARS-CoV-2 infection, negatively impacting the control of virus spread. Many alternatives have enabled the continuous processing of samples and have presented a decrease in time and cost. These measures thus allow broad testing of the population and should be ideal for controlling the disease. In this sense, we compared the SARS-CoV-2 molecular detection effectiveness by Real time RT-PCR using two different protocols for RNA extraction. The experiments were conducted in the National Institute of Health (INS) from Peru. We compared Ct values average (experimental triplicate) results from two different targets, a viral and internal control. All samples were extracted in parallel using a commercial kit and our alternative protocol-samples submitted to proteinase K treatment (3 μg/μL, 56°C for 10 minutes) followed by thermal shock (98°C for 5 minutes followed by 4°C for 2 minutes); the agreement between results was 100% in the samples tested. In addition, we compared the COVID-19 positivity between six epidemiological weeks: the initial two in that the Real time RT-PCR reactions were conducted using RNA extracted by commercial kit, followed by two other using RNA obtained by our kit-free method, and the last two using kit once again; they did not differ significantly. We concluded that our in-house method is an easy, fast, and cost-effective alternative method for extracting RNA and conducing molecular diagnosis of COVID-19. |
| format | Article |
| id | doaj-art-71dd41c43bef470b8eea72e8f34a6810 |
| institution | Kabale University |
| issn | 1932-6203 |
| language | English |
| publishDate | 2021-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj-art-71dd41c43bef470b8eea72e8f34a68102025-08-20T03:46:20ZengPublic Library of Science (PLoS)PLoS ONE1932-62032021-01-01163e024888510.1371/journal.pone.0248885A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock.Adolfo Marcelo ÑiqueFiorella Coronado-MarquinaJairo Andrés Mendez RicoMaría Paquita García MendozaNancy Rojas-SerranoPaulo Vitor Marques SimasCesar Cabezas SanchezJan Felix DrexlerOne of the biggest challenges during the pandemic has been obtaining and maintaining critical material to conduct the increasing demand for molecular tests. Sometimes, the lack of suppliers and the global shortage of these reagents, a consequence of the high demand, make it difficult to detect and diagnose patients with suspected SARS-CoV-2 infection, negatively impacting the control of virus spread. Many alternatives have enabled the continuous processing of samples and have presented a decrease in time and cost. These measures thus allow broad testing of the population and should be ideal for controlling the disease. In this sense, we compared the SARS-CoV-2 molecular detection effectiveness by Real time RT-PCR using two different protocols for RNA extraction. The experiments were conducted in the National Institute of Health (INS) from Peru. We compared Ct values average (experimental triplicate) results from two different targets, a viral and internal control. All samples were extracted in parallel using a commercial kit and our alternative protocol-samples submitted to proteinase K treatment (3 μg/μL, 56°C for 10 minutes) followed by thermal shock (98°C for 5 minutes followed by 4°C for 2 minutes); the agreement between results was 100% in the samples tested. In addition, we compared the COVID-19 positivity between six epidemiological weeks: the initial two in that the Real time RT-PCR reactions were conducted using RNA extracted by commercial kit, followed by two other using RNA obtained by our kit-free method, and the last two using kit once again; they did not differ significantly. We concluded that our in-house method is an easy, fast, and cost-effective alternative method for extracting RNA and conducing molecular diagnosis of COVID-19.https://doi.org/10.1371/journal.pone.0248885 |
| spellingShingle | Adolfo Marcelo Ñique Fiorella Coronado-Marquina Jairo Andrés Mendez Rico María Paquita García Mendoza Nancy Rojas-Serrano Paulo Vitor Marques Simas Cesar Cabezas Sanchez Jan Felix Drexler A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock. PLoS ONE |
| title | A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock. |
| title_full | A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock. |
| title_fullStr | A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock. |
| title_full_unstemmed | A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock. |
| title_short | A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock. |
| title_sort | faster and less costly alternative for rna extraction of sars cov 2 using proteinase k treatment followed by thermal shock |
| url | https://doi.org/10.1371/journal.pone.0248885 |
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