The present study analyzed tumor protein p53 gene (TP53) polymorphism as a factor in uterine implantation and establishment of pregnancy in four old-aged Japanese Black cows with long-term infertility. They had G allele or A allele in exon 6 of the gene. Oocytes were collected from these females using ovum pick up (OPU) and fertilized in vitro. Only some of the oocytes from individuals having G allele had developmental competence to the blastocyst stage, while no oocytes having the A allele had the competence to develop to the blastocyst stage. In conclusion, we suggest that a cause of long-term infertility in old-aged Japanese Black cows may be a polymorphism of TP53 affecting the competence of embryonic development.