Abstract Background Detection of SARS-CoV-2 viral RNA is important for the diagnosis and management of COVID-19. Methods We present a clinical validation of a RT-PCR assay for the SARS-CoV-2 nucleocapsid (N1) gene. Offboard lysis on an automated nucleic acid extraction system (EMAG®) was optimized with endemic Coronaviruses (OC43 and NL63). Genomic RNA and SARS-CoV-2 RNA in a recombinant viral protein coat (Accuplex) were used as control materials and compared for recovery from nucleic acid extraction. Results Nucleic acid extraction showed decreased recovery of endemic Coronavirus in vitro transcribed RNA (NL63) compared to attenuated virus (OC43). SARS-CoV-2 RNA (Accuplex) had more reliable recovery from extraction through amplification compared to genomic RNA. Recovery of genomic RNA was improved by combining lysis buffer with clinical matrix prior to adding RNA. The RT-PCR assay demonstrated 100% in silico sensitivity and specificity. The accuracy across samples was 100% (75 of 75). Precision studies showed 100% intra-run, inter-run, and inter-technologist concordance. The limit of detection was 264 copies per ml (estimated 5 copies per reaction; 35.56 mean Ct value). Conclusions This SARS-CoV-2 assay demonstrates appropriate characteristics for use under an emergency use authorization. Endemic Coronavirus controls were useful in optimizing the extraction procedure. In the absence of live or attenuated virus, recombinant virus in a protein coat is an appropriate control specimen type for assay validation during a pandemic.