App Note
Extraction of SARS-Cov-2 virus from the supernatant of infected Vero E6 cells using RNAGEM V
Coronaviruses (CoVs) are a group of RNA viruses able to cause infections primarily in birds and mammals. In 1960, it was discovered that coronaviruses can also infect humans, causing mild to severe respiratory infections. Particularly, in the last two decades, these viruses have provoked lethal
diseases in humans such as SARS in 2003, MERS in 2012, and COVID-19 in 2019.
COVID-19 was identified for the first time in Wuhan, China, in December 2019, and it is a disease of the respiratory tract caused by a novel coronavirus named SARS-CoV-2. In contrast to SARS and MERS, SARS-CoV-2 has spread faster leading to a pandemic situation that counts globally more than
25,000,000 of infected people and more than 850.000 deaths as of September 2020, according to Coronavirus Resource Center at Johns Hopkins University.
In absence of a specific therapy, both early diagnostic and monitoring at population level have been proven to be the most effective method to counteract the spread of COVID-19.
The gold standard for early detection of SARS-CoV-2 worldwide is RT-qPCR. Therefore, the RNA extraction represents a key pre-analytical step. However, the large number of diagnostic tests required caused a shortage of reagents, including RNA extraction kits. In addition, conventional RNA extraction
methods such as columned and beads-based can be costly and time consuming. Therefore they are not always adequate to respond timely to the high-throughput extraction required during this COVID-19 pandemic.
In order to provide a fast and cost-effective RNA extraction method, we tested whether RNAGEM V can extract rapidly viral RNA from SARS-CoV-2 grown in cultured Vero E6 cells. RNAGEM V (#RTV1000) is a Research Use Only (RUO) product that allows a simple and rapid RNA extraction
leveraged by a temperature-driven, single-tube approach. Activated at 75°C and inactivated at 95°C, this broad specificity, thermophilic proteinase simultaneously lyses cells and eliminates RNases, thereby stabilizing the RNA and maintaining its integrity. The single-tube extraction protects
precious samples, ensuring all RNA is preserved. In addition, RNAGEM V has already been utilized for research use to extract viral RNA SARS-COV-2 collected from nasopharyngeal swab and analysed via microarray in comparison to the traditional spin-column extraction and analysis via qRT-PCR
(Damin et al. 2021). Besides SARS-COV-2, RNAGEM V has been successfully utilized to extract viral RNA fromfrom other viruses such as Canine distemper virus (CDV; DFE-CE strain), HCV, Equine H3N8, and H7N7 viruses. In agreement with the successful extraction of viral RNA from the mentioned viruses, RNAGEM V was able in this work to extract RNA from SARS-CoV-2.
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RNAGEM V
Effortless, simultaneous nucleic acid extraction from DNA and RNA viruses … in minutes, not hours.
RNAGEM V uses MicroGEM’s novel thermophilic proteinase and buffer system to extract nucleic acids from viruses in less than 15 minutes without the need for purification.
Sample types: nasal-pharyngeal swabs in saline solution, nasal swabs, buccal swabs, and sputum/saliva
Downstream applications: PCR, RT-PCR, RT-qPCR, LAMP, RT-LAMP
Key Advantages:
- Nucleic acid extraction in less than 15 minutes
- Simultaneous extraction from DNA and RNA viruses
- Single tube extraction
- No ionic detergents or chaotropic salts
- No magnetic beads – No spin columns
- High nucleic acid recovery – Minimal loss of nucleic acids during extraction
- Flexibility – suitable for low-throughput to high-throughput extraction with a single protocol
- Easily automated using standard liquid handling solutions
- Minimal plasticware required – Reduced waste and supply chain issues
Product Overview (pdf)
Quick Start Guide (pdf)
RNAGEM V is for Research Use Only (RUO) and is not intended for in vitro diagnostic (IVD) use.
RNAGEM
Temperature-driven, single-tube extractions of total RNA from mammalian cells, tissues, insects, bacteria and virus
This powerful, broad-specificity approach is ideal for gene expression from a very small number of cells, such as exosomes or single cells, without the need for harsh chemicals or further purification.
- Ideal for preparing RNA from mammalian cell culture, laser capture microdissection, FACS-prepared cell populations, cells in suspension, adherent cells, cells stored in RNAlaterTM and cell pellets
- Simplified, hands-off workflow provides RT-PCR and RT-qPCR ready RNA in approximately 5 minutes
- Releases both RNA and DNA with excellent recovery across a wide range of cell numbers
- Requires no further purification for accurate RT-PCR and RT-qPCR analysis
- Inhibitor-free means no harsh chemical washes or multiple steps
- RT-PCR and RT-qPCR reagents can be added directly to the lysate for a simple, streamlined protocol in 96 well plates
- Each kit contains RNAGEM, DNase I, BLUE buffer, 10x DNase buffer, and 10x TE
Learn more about RNAGEM and recent applications. Download our RNAGEM technical overview. Watch our webinar on advances in extracting and stabilizing RNA.