Development of canine X-chromosome inactivation pattern analysis for the detection of cell clonality by incorporating the examination of the SLIT and NTRK-like family member 4 (SLITRK4) gene

X-chromosome inactivation pattern (XCIP) analysis can be used to assess the clonality of cell populations of various origin by distinguishing the methylated X chromosome from the unmethylated X chromosome. In this study, the utility of XCIP analysis was improved by incorporating the examination of AC dinucleotide repeats in SLIT and NTRK-like family member 4 (SLITRK4) gene into the previously reported CAG repeat examination of androgen receptor (AR) gene in dogs. The rate of heterozygosity when both genes were analysed (125/150, 83.3%) was higher than AR gene examination alone (86/150, 57.3%). Blood samples from heterozygous dogs in either AC-1 or AC-2 of SLITRK4 gene were examined for the corrected inactivation allele ratio (CIAR), resulting in the determination of a reference range of CIAR < 3.8 in non-neoplastic cell/tissue samples. Using this analytical method, 49% (21/43) of neoplastic tissue samples from dogs showed a CIAR > 3.8, indicating the presence of a clonal population. Through the present study, the availability of canine XCIP analysis was improved by incorporating the examination of the SLITRK4 gene, providing a highly useful laboratory examination system for the detection of the clonality of various cell/tissue samples in dogs.