RhD-negative blood exhibits various phenotypes, each having multiple subtypes, and all associated with specific molecular mechanisms. This study reports a case involving a nucleotide variation at position 767 in exon 5 of the RhD gene (c.767C>A, p.Ser256Ter), resulting in the substitution of serine with a termination codon at position 256. These changes led to structural alterations in the RhD antigen protein, resulting in a serologically RhD-negative phenotype in the patient. The sequence data of this allele was reported for the first time and has been submitted to GenBank under the accession number PQ740962. For RhD-negative patients, serological and molecular biological methods should be further used to determine their subtypes and molecular genetic background. Different RhD blood types require differential management for pregnant women, transfusion recipients, and blood donors. Therefore, it is necessary to obtain more information for prenatal monitoring and transfusion management. This novel RhD allele mutation enriches the understanding of the molecular biological mechanisms underlying the formation of RhD-negative blood. It contributes to the accurate determination of RhD blood type and the development of "precision" blood transfusion guidance for clinical practice, thereby reducing transfusion risks for patients with rare blood types and ensuring transfusion safety.