The microvirid phage φK, specific for Escherichia coli K12, contains a circular singlestranded (SS) DNA in the icosahedral virion, which comprises four phage gene products, F (capsid), G (major spike), H (minor spike), and J (core). φKhT, a host-range mutant of φK, can grow on E. coli C and B, besides K12, and is more thermosensitive than the parental phage φK. Sequencing analysis revealed that the genome of φK and φKhT consists of 6, 089 nucleotides (nt), and codes for eleven genes, whose sequences are similar to those of α3, φX174, and G4 infective to strain C. In φKhT, two nt had changed: one is in the gene G, resulting in replacement of the 75th codon Ala with Ser, and the other is at 67th codon of the gene H: Val to Ala. Chemically synthesized gene J protein composed of 23 amino acids (aa) binds to φK SS DNA more tightly than and preferentially over the host E. coli SS-DNA-binding protein (SSB). These results indicate that the two spike proteins G and H are involved in the determination of φK host-range, and support a model in which the gene J protein functions in packaging the viral SS DNA into the virion vesicle.