Abstract Crossbreeding using exotic bulls have been developed widely in Indonesia. Y-chromosome markers have high value for the evaluation of paternal genetic and phylogeny history. The aim of this study was to investigate the coding sequence polymorphism of SRY gene in crossbred cattle using Belgian Blue Bull and Wagyu Bull. Polymerase chain reaction method was used to identify the target sequence 690 bp of SRY gene coding sequences. Then, direct DNA sequencing with forward primer was performed to identify the single nucleotide polymorphism (SNP) differences between individual sample. The results of the alignment coding sequence of SRY gene from four GenBank data, Brahman cross cattle, Belgian Blue Bull Cross and Wagyu Bull cross revealed that there are 21 SNPs variation. Six SNPs out of 21 SNPs (29%) was synonymous and 15 SNPs out of 21 SNPs (71%) was change the amino acid (non-synonymous). Eight out of 21 SNPs (38%) were transition mutation and 13 SNPs out of 21 SNPs (62%) were transversion mutation. Crossbred cattle using Brahman cross cow with Belgian Blue Bull and Wagyu Bull show the SNP at position 1707T→G (transversion mutation). This SNP change the amino acid from Phe(TTT) to Cys(TGT). Thus, the result can be used as molecular marker for identifying the paternal lineage of crossbred cattle which recently develop in Indonesia.