A library of overlapping cDNAs obtained from Norwalk virus purified from stools of human volunteers (Jiang et al., 1990, Science 250, 1580-1583) was used to obtain the nucleotide sequence of the viral genome. The sequence has a total of 7642 nucleotides, excluding the 3' poly(A) tail, and has a base composition of 48% G + C. Three open reading frames (ORF) are predicted in the sequence. The longest ORF (ORF1, nucleotides (nt) 146 to 5359) is predicted to encode a polyprotein precursor to nonstructural proteins based on identification of sequences similar to the picornavirus 2C protein, 3C protease, and 3D RNA-dependent RNA polymerase. ORF2 (nt 5346 to 6935) is predicted to encode a polypeptide with a predicted molecular weight of 56,571 (56.6K, close to the expected size of the viral capsid protein), and it contains a short region of sequence similarity to the picornavirus structural protein VP3. A third potential ORF (nt 6938 to 7573) could encode a small polypeptide of 22.5K. The genomic organization found in Norwalk virus shares striking similarities with the genome of two caliciviruses, the feline calicivirus and the rabbit hemorrhagic disease virus. The morphology, size, polarity, and genomic organization of the Norwalk virus indicate it is a member of the Caliciviridae family.