The spread of 2019 novel coronavirus (2019-nCoV) poses a serious threat to human health. We report here the isolation of a 2019-nCoV strain named nCoV-SH01 from the throat swab from a COVID-19 patient in Shanghai. The isolate grows well on Vero E6 cells with typical syncytial cytopathic effects. The nCoV-SH01 genome has been sequenced using sanger sequencing and Illumina sequencing and analyzed, which shares more than 99.99% identity to a reference sequence (GenBank MN988668 2019-nCoV Wuhan-Hu-1 isolate) with a difference of only three nucleotides. Immunofluorescence results showed that nCoV-SH01-infected cells reacted with the serum of a convalescent COVID-19 patient, suggesting virus replication in cultured cells. The isolation of nCoV-SH01 provides the basis on which cell and animal infection models would be established to facilitate the study of 2019-nCoV pathogenicity and the development of antiviral drugs and vaccines.