Homemade viral RNA isolation protocol using silica columns: A comparison of four protocols

Gonzalez-Perez, Idania; Cayarga, Anny Armas; Rosa, Iria García de la; González, Yaimé Josefina González

doi:10.1016/j.ab.2006.10.022

Cited by 6 publications

(5 citation statements)

References 6 publications

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“…However, the extracted RNA from this method is suitable for HCV molecular diagnosis by real-time RT-PCR, and by RT-LAMP when using samples with a high viral load. Previous studies also showed that this method was suitable for RNA isolation from plasma and ready for use in subsequent reactions for viral detection [ 7 , 8 ]. The spin-column-based method also efficiently extracted DNA and facilitated the amplification of targets by LAMP and PCR [ 9 ].…”

Section: Discussionmentioning

confidence: 99%

Comparison of Simple RNA Extraction Methods for Molecular Diagnosis of Hepatitis C Virus in Plasma

et al. 2022

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Nucleic acid extraction from biological samples is an important step for hepatitis C virus (HCV) diagnosis. However, such extractions are mostly based on silica-based column methodologies, which may limit their application for on-site diagnosis. A simple, rapid, and field-deployable method for RNA extraction is still needed. In this study, we evaluated the efficacy of four simple RNA extraction methods for the detection of HCV in plasma samples: a silica-membrane-based method, a magnetic-beads-based method, boiling with diethyl pyrocarbonate (DEPC)-treated distilled water, and using a commercial lysis buffer. HCV RNA was detected using both real-time reverse transcription polymerase chain reaction (RT-PCR) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). Using real-time RT-PCR, extracted RNA from the silica-membrane-based and magnetic-beads-based methods had a 100% detection rate for RNA extraction from plasma. Using RT-LAMP, extracted RNA from the silica-membrane-based method showed a 66% detection rate, while the magnetic-beads-based method had a 62% detection rate. In summary, magnetic-beads-based extraction can be used as an alternative RNA extraction method for on-site HCV detection. Boiling with DEPC-treated distilled water was not appropriate for low HCV load samples, and boiling with a lysis buffer was not recommended.

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Section: Discussionmentioning

confidence: 99%

Comparison of Simple RNA Extraction Methods for Molecular Diagnosis of Hepatitis C Virus in Plasma

et al. 2022

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“…However, the extracted RNA from this method is suitable for HCV molecular diagnosis by real time RT-PCR, and by RT-LAMP when using samples with a high viral load. Previous studies also showed that this method was suitable for RNA isolation from plasma and ready-for-use in subsequent reactions for viral detection [7, 8]. This extraction method is fast but the most expensive of the four extraction methods tested.…”

Section: Discussionmentioning

confidence: 90%

Comparison of simple RNA extraction methods for molecular diagnosis of hepatitis C virus in plasma

Hongjaisee

Jabjainai

Sakset

et al. 2022

Preprint

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We evaluated the efficacy of four simple RNA extraction methods for the detection of hepatitis C virus (HCV) in plasma samples: silica-membrane-based, magnetic beads-based, boiling with Diethyl Pyrocarbonate (DEPC)-treated distilled water or a commercial lysis buffer. HCV RNA was detected using both real time reverse transcription polymerase chain reaction (RT-PCR) and reverse transcription loop mediated isothermal amplification (RT-LAMP). The magnetic beads-based extraction can be used as an alternative RNA extraction method for on-site HCV detection. Boiling with DEPC-treated distilled water was not appropriate for low HCV load samples and boiling with a lysis buffer was not recommended.

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“…However, this is such a vital step in the extraction procedure that without proper calibration, standardization, and sterility, large variations in yield and purity are likely to compromise the results. There have been previous attempts to create silica-based extraction protocols that rely on in-house buffers and commercially available columns [11], yet the use of phenol and other hazardous reagents in these protocols can become a problem if not handled properly and could result in injury if the user is still new to the protocol. Although guanidine thiocyanate is minimally hazardous, it is considerably less hazardous than the aforementioned phenol and chloroform.…”

Section: Resultsmentioning

confidence: 99%

A cost-effective RNA extraction technique from animal cells and tissue using silica columns

Escobar

Hunt

2017

J Biol Methods

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Ribonucleic acid (RNA) is widely used in molecular biology assays, and some of the most common assays include: northern blotting and RT-PCR gene expression analysis. RNA is generally extracted by two methods: phenol-chloroform or commercially available silica spin column kits. Phenol-chloroform extraction is generally more economical; however, it produces hazardous byproducts, and leftover chemicals in the sample that can inhibit downstream applications. Commercial kits usually have simple set ups and short preparation time; however, they can introduce a significant expense to laboratory budgets. Here we have created a method to extract RNA using generic silica columns and readily available reagents while maintaining a high yield and purity.

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Homemade viral RNA isolation protocol using silica columns: A comparison of four protocols

Cited by 6 publications

References 6 publications

Comparison of Simple RNA Extraction Methods for Molecular Diagnosis of Hepatitis C Virus in Plasma

Comparison of Simple RNA Extraction Methods for Molecular Diagnosis of Hepatitis C Virus in Plasma

Comparison of simple RNA extraction methods for molecular diagnosis of hepatitis C virus in plasma

A cost-effective RNA extraction technique from animal cells and tissue using silica columns

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