SARS-CoV-2 ( Severe acute respiratory syndrome coronavirus) is a causative agent of Coronavirus disease 2019 or COVID -19 is a viral disease whereas the provisional name 2019 novel coronavirus (2019-nCoV). It is a positive-sense single-stranded RNA virus and strain was first discovered in Wuhan, China, so it is sometimes referred to as the “Wuhan virus” or “Wuhan coronavirus”. Classification of this virus is as-
Realm: Riboviria
Kingdom: Orthornavirae
Phylum: Pisuviricota
Order: Nidovirales
Family: Coronaviridae
Genus: Betacoronavirus
Subgenus: Sarbecovirus
Species: Severe acute respiratory syndrome-related coronavirus
Strain: Severe acute respiratory syndrome coronavirus 2
Primary reservoir bat whereas an intermediate animal reservoir such as a pangolin is also thought to be involved in its introduction to humans. The virus is primarily spread between people through close contact and via respiratory droplets produced from coughs or sneezes. It mainly enters human cells by binding to the receptor angiotensin-converting enzyme 2 (ACE2).
RNA is a polymeric substance present in SARS-CoV-2, consisting of a long single-stranded chain of phosphate and ribose units with the nitrogen bases adenine, guanine, cytosine, and uracil, which are bonded to the ribose sugar. RNA is used in all the steps of protein synthesis as well as carries the genetic information in this virus.
The isolation of RNA with high quality is a crucial step required to perform various molecular experiments. It is based on silica gel purification. The sample is lysed and digested with lysate and protease, RNA is released into the lysate. Transfer to an adsorption plate and filter column. RNA is adsorbed on the membrane, while protein is not adsorbed and is removed with filtration.
After washing proteins and other impurities are washed and RNA is finally eluted with low-salt buffer ( 10 MmTris, pH 8.0).
Hipure viral mini-column ( Adsorption column)
PK/Carrier RNA (Proteinase/Poly A)
Protease Dissolve buffer( Glycerol/Tris/CaCl2)
Buffer VLF (Guanidine salt/ isopropanol)
Buffer CW (Tris/NaCl/Isopropanol)
Buffer AVE (Tris, pH 8.0)
Biosafety cabinet 2A extraction room ( due to being an organism, SARS-CoV-2 of risk group 3)
Specimen Collection Methods
Collect specimen in virus transport medium (VTM).
Arrange for transportation in a way the sample is received within 24 hours.
Store collected specimen at 2 – 4° C (refrigerator) till ready for transportation.
NEVER STORE THE SAMPLE AT FREEZER COMPARTMENT(-20° C) as it may degrade the sample quality.
Transportation Of Specimen
Transport specimen in 3 layers packaging system maintaining cold chain (2-4° C). Submit specimen with appropriate form fill up issued by WHO inappropriate testing site.
Sample Packaging for Coronavirus disease (COVID -19)
Layer 1: Put VTM with the specimen in a zip lock bag.
Layer 2: Place the zip lock bag in an airtight container lined with absorbent material(cotton).
Layer-3: Place this air-tight container in an icebox/vaccine box with an ice pack in all four corners as shown above image.
Sample rejection criteria if any
The sample is transported without maintaining a cold chain.
The sample is received without proper form fill up.
Unlabeled sample.
10 µl
1000 µl (100-1000 µl adjustable)
40 µl