Date of Award
Master of Science
Clinical Molecular Genetics
A novel coronavirus called SARS-CoV-2 has caused the emergency release of the most well-known molecular assay, the CDC N1 RT-PCR assay, causing reports of poor analytical performance resulting in false-negative results (20, 26), and inconsistent testing kits received (23, 32). An immediate and dire need for a rapid and reliable SARS-CoV-2 testing workflow specifically designed for a university setting is the purpose this project is aiming and intended to fulfill. The workflow design uses a less invasive saliva sample for rapid screening using colorimetric RT-LAMP detection of three SARS-CoV-2 gene regions for Orf1ab, envelope, and nucleocapsid. Purified SARS-CoV-2 genomic RNA is spiked into Proteinase K and heat-treated saliva from a SARS-CoV-2 negative donor to simulate a positive donor sample used to characterize and optimize this workflow, detecting 200 copies of SARS-CoV-2 genomic RNA by all three gene targets. The utility of colorimetric RT-LAMP outperformed the CDC N1 RT-PCR assay in turn-around-time and analytical performance. When applied to a small surveillance study, five out of 47 asymptomatic saliva donors had (September 2020) detectable SARS-CoV-2 genetic material, resulting in a 10% positivity rate, with the campus dashboard reporting
Juntila, Casey, "SENSITIVE AND SPECIFIC DETECTION OF SARS-COV-2 IN SALIVA USING REVERSE TRANSCRIPTASE LOOP-MEDIATED ISOTHERMAL AMPLIFICATION" (2021). All NMU Master's Theses. 679.