Abstract: Objective To investigate the RHD genetic background of 48 D variant specimens. Methods Specimens were collected from blood donors and patients sent to Suzhou Blood Center and Xuzhou Blood Center for RHD blood group confirmation from July 2020 to July 2021. All samples showing negative or equivocal agglutination were tested by IAT with three commercially available anti-D reagents,with D variant defined once it reacted positively with an anti-D. The genomic DNA of the D variant specimen was extracted,and the RHD genotype of the specimen was preliminarily identified by PCR-SSP. For doubtful or unidentifiable results,further genetic sequencing of RHD exons 1-10 was performed. Results A total of 48 cases were identified with the D variant.The PCR-SSP method directly determined the allelic composition in 35 cases, including RHD*15,RHD*06.03.01,RHD*15/RHD*01EL.01 and RHD*01EL.01,of which weak D type and DⅥ type are the most prevalent,with 17 cases（34.69%）and 9 cases（18.37%）respectively. The allelic composition of the remaining 13 cases was determined after RHD sequencing,including 5 cases of weak D（RHD*01W.72,RHD*01W.54, RHD*01W.10 and RHD*11/RHD*01EL.01）,4 cases of partial D（RHD*05.01,RHD*05.04,RHD*05.05 and RHD*16.02）,as well as 1 case of RHD*01. There were also 3 weak D specimens with single nucleotide mutations,2 cases with RHD*779G and 1 case with 509T>G. The 509T>G mutation was found to be like partial D after being tested with multiple monoclonal antibodies. The common type of RHD*15 was ccEe,accounting for 70.59% of the allelic composition; the common type of RHD*06.03.01 was Ccee,accounting for 77.78% of the allelic composition. Conclusion RHD*15 is the most common D variant in the two areas,whose common type is ccEe,and the D variant has rich genetic polymorphisms for this locus.

Key words: D variant, RHD gene, RHD negative