The Dutt lab at ACTREC-Tata Memorial Centre developed a real-time RT PCR-based one-step, one-tube, multiplex diagnostic kit to detect the COVID-19 virus. The kit was successfully transferred to industry through a formal institutional MOU. It’s the first diagnostic kit from an Indian academic institute with peer-reviewed methodology, published in Heliyon. In brief, the group identified a unique conserved region in the SARS-CoV-2 Nucleocapsid gene. The fragments were cloned for generating synthetic RNA molecules for three such regions (N1, N2, and N3) and spiked in the human genomic DNA and total RNA at varying concentrations as a reference sample to mimic a COVID-19 positive sample. The reference sample was used to compare the detection limit of two different chemistries: a probe-free SYBR Green dye-based methodology and TaqMan probe-based RT-PCR assay. The probe-based assay was found to be more sensitive with detection of up to 15 RNA copies, as compared to 150 RNA copies of the probe-free method. This was further developed into a one-step, one-tube, multiplex diagnostic kit. The detection accuracy of the kit was evaluated using a panel of clinical specimens by performing over 200 validations with dilutions of samples ranging from 1-100 ng. The kit was 100% specific and 100% sensitive based on the accurate identification of clinical samples. Furthermore, to minimize variability and automate the qRT-PCR downstream analysis of multiple samples in the same run, we developed an open-access freely available GUI based analytical tool, COVID qPCR Analyzer tool (www.actrec.gov.in/pi-webpages/AmitDutt/Covid/Analyzer.html), and in silico automated tool to detect the virus in an unambiguous and reproducible manner with minimal requirement of third-party tools in a platform-independent for its universal application across all kinds of real-time PCR machine. Several labs, including the Suraksha Diagnostic Pvt Ltd, Genetic Lab SRL, and Genes N Life Health Care Pvt Ltd., and the Univ of British Columbia, Canada The Univ Hospital of La Réunion, France, for academic purpose, have downloaded this tool.

​

​[Sci Soup] [The Hawk] [Biotech Times] [India Today] [Research Stash]