Cold-adapted influenza viruses A/Leningrad/13 4/17/5 7 (H2N2) (Len/17) and A/Leningrad/I 34/4 7/57 (H2N2) (Len/47) are used in Russia to prepare live reassortant cold-adapted influenza vaccines (LIV) for adults and children, respectively. Comparison between the nucleotide sequences of the Len/17 strain and the initial wild-type strain A/Leningrad/13 4/5 7 (H2N2) revealed ten nucleotide substitutions (eight of them encoding). Four additional substitutions (three encoding) were found in the genome of the Len/47 virus. Gene segment restriction site (PCR-restriction) analysis was used for identification of the genotype of reassortant influenza viruses. Conventional methods of PCR-restriction analysis detect only five encoding nucleotides substitutions in the internal genes of the Len/17 and seven substitutions in the internal genes of the Len/47 virus. An extended modification of the PCR-restriction method detect all encoding mutations in the internal genes of the Len/17 and Len/47 viruses (eight and eleven encoding substitutions, respectively). This method is advantageous for genome composition analysis of reassortant influenza vaccine strains and for investigating the genetic stability of LIV during replication in vaccines.