This study was conducted to isolate Shigella SPP. from (150) stool samples of children with both sex suffering from diarrhoea whose ages ranged from (1-60) months during the period from October 2021 to March 2022 in Mosul / Iraq. The results showed that (17) isolates belong to Shigella sonnei, representing (12.59%) of the total number of isolates, The age group (49-60) months was in isolating to Shigella sonnei which diagnosis by morphological and biochemical. The sequence analysis of the DNA was carried out, especially for the ribosomal 16SrRNA gene, it was relied upon because it is a fixed sequence that does not change over the years for bacterial species and genera. The DNA extract was sent to the Microgene Company to target the diagnostic gene 16SrRNA by adding the special Forward and Reverse primer genes and amplification and doubling polymerase chain reaction (PCR), It was found the sequence of gene for all the (17) samples was present in all strains. When comparing the duplicated bands, it was found that the bands were (1460) base pairs in size and at the same level for all types of strains. The results of these strains were compared for the genetic sequence with the gene sequence of the standard strains installed in the database Basic global Alignment Search Tool (BLAST) within National Center for Biotechnology Information (NCBI). The diagnosis was confirmed with a cause similarity ranging from (98- 100%).Found that the size of the nitrogenous bases reaches (1460) base pairs when electrophoresis after 16SrRNA gene amplification when electroporation after 16SrRNA gene amplification by polymerase chain reaction (PCR). Conclusion: Bacteria were diagnosed by PCR as a confirm test for Shigella sonnei.