The recent pandemic outbreak of COVID-19 caused by a novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), poses a threat to public health globally. Thus, developing a rapid, accurate, and easy-to-implement diagnostic system for SARS-CoV-2 is crucial for controlling infection sources and monitoring illness progression. Here, we reported an ultrasensitive electrochemical detection technology using calixarene functionalized graphene oxide for targeting RNA of SARS-CoV-2. Based on a supersandwich-type recognition strategy, the technology was confirmed to practicably detect the RNA of SARS-CoV-2 without nucleic acid amplification and reverse-transcription by using a portable electrochemical smartphone. The biosensor showed high specificity and selectivity during analysis and actual testing. A total of 88 RNA extracts from 25 SARS-CoV-2-confirmed patients and eight recovery patients were detected using the biosensor. The detectable ratios (85.5 % and 46.2 %) were higher than those obtained using RT-qPCR (56.5 % and 7.7 %). The limit of detection (LOD) of the clinical specimen was 200 copies/mL, which is the lowest LOD among the published RNA measurement of SARS-CoV-2 to date. Additionally, only two copies (10 μL) of SARS-CoV-2 were required for per assay. Therefore, we developed an ultrasensitive, accurate, and convenient assay for SARS-CoV-2 detection, providing a potential method for point-of-care testing.