RHD Gene Analysis of A Blood Donor with Del Phenotype / 中国实验血液学杂志

OBJECTIVE@#To analyze the RHD genotype of a blood donor with Del phenotype in Yunnan.@*METHODS@#Rh serological phenotype was identified. RHD gene was detected by PCR-SSP typing, and its 10 exons were sequenced. Exon 9 was amplified for sequencing and analysis. RHD zygosity was detected.@*RESULTS@#The Rh phenotype of this specimen was CcDelee. Genomic DNA exhibited a 1 003 bp deletion spanning from intron 8, across exon 9 into intron 9. The deletion breakpoints occurred between two 7-bp short tandem repeat sequences. There was no variation in the sequences of the remaining exons. The Rh hybridization box test showed that there was one RHD negative allele.@*CONCLUSION@#This specimen is Del type caused by deletion of RHD exon 9.

Study on RHD and RHCE gene mRNA based on three generation sequencing technology / 中国输血杂志

【Objective】 To establish RH gene mRNA sequencing method based on nanopores sequencing and to explore the RHD and RHCE mRNA transcripts in D positive and Del individuals. 【Methods】 From March 2021 to May 2022, 5 RhD positive samples and 5 Del samples screened out by hospitals in Chengdu were sent to our laboratory for futher examination. The erythrocytes and buff coat were isolated, then DNA and RNA were extracted.All 10 samples were genotyped by PCR-SSP. After the mRNA was reversely transcribed into cDNA, the full-length mRNA of RHD and RHCE genes were simultaneously amplified by a pair of primers. Sanger sequencing and third-generation sequencing technology based on Nanopore were used to sequence the amplified products, and the types and expressions of different splices of RHD and RHCE gene mRNA transcripts were analyzed. 【Results】 The method established in this study can simultaneously amplify the full length transcripts of RHD and RHCE. Ten different RHD gene mRNA transcripts and nine RHCE gene mRNA transcripts were detected in 10 samples. RHD full-length transcript (RHD-201) can be detected in RhD Del type, but the expression amount was significantly lower than that in RhD positive samples. The expression amount of transcript RHD-207 (Del789) in Del samples was significantly higher than that in RhD positive samples. The transcript RHD-208 (Del8910+ 213) was only detected in RhD Del type individuals, and no significant difference was found between other RHD transcripts and all RHCE transcripts in the two phenotypes. 【Conclusion】 In this study, an analytical method for sequencing full-length transcript isomers of RHD and RHCE mRNA via the third generation was successfully established, and complex alternative splicing patterns were found in RHD and RHCE genes, providing a new method for the study of alternative splicing of blood group gene variants mRNA.

Molecular basis of Del phenotype in RhD negative population of Zhongshan / 中国输血杂志

【Objective】 To conduct serological and molecular study of Del type in RhD-negative donor population in Zhongshan area, so as to improve the diagnosis of Del type. 【Methods】 A total of 102 initially RhD-negative samples, collected from December 2017 to February 2019, were classified by RHCE and PCR-SSP genotyping. And 95 cases of truly negative RhD were confirmed by IAT, 28 cases of Del type were identified by absorption and elution test. The phenotype and genotyping characteristics of Del type in Zhongshan area were summarized based on domestic data of relative literature. 【Results】 Among 102 initially RhD-negative samples by serological test, 95 were truely RhD-negative, 28 were DELRHD 1227A without any other Del allele. Among them, RHCE antigen type were Ccee in 20(71.4%) cases, CCee in 8(28.6%), with no difference in comparison with other regions in China. The frequency of Del in RhD-negative blood donors was 29.5% (28/95), with difference between Shanghai, Taiwan, and Fuzhou, but no difference between Nanchang, Zhejiang, and Wuhan. 【Conclusion】 The study showed that the Del phenotype was closely related to Ce haplotype, and has no difference with other regions in China. The frequency of Del type in RhD negative donors was 29.5%, with regional differences. RHD1227A was the main allele of Del.

Frequency analysis of RHD*1227A allele among Rh negative Han population and random population in Liaoning / 中国输血杂志

【Objective】 To investigate the frequency of RHD*1227A allele in Rh negative Han polulation and random population in Liaoning. 【Methods】 Del phenotype was screened by absorption-elution test, the RHD*1227A allele was screened by PCR-SSP and confirmed by nucleotide sequence analysis of RHD full coding region, and the heterozygosity of RHD gene was detected by hybridization technique. 【Results】 24 case of Del phenotype were detected by the absorption-elution test among 117 Rh negative individuals. 23 RHD*1227A allele carriers were detected by PCR-SSP and sequencing-based typing (SBT). Genotype of 19 individuals was RHD*1227A/RHD*01N.01, while the other 4 was homozygous RHD*1227A/ RHD*1227A.11 individuals were detected as RHD*1227A allele among 1 045 random blood donors, among which 9 were RHD*1227A/RHD*01 and 2 were RHD*1227A/RHD*01N.01. 【Conclusion】 The frequency of RHD*1227A allele is 0.115 4 in Rh negative Han population and 0.005 3 in random population in Liaoning..

Investigation on the distribution of weak D and Del phenotype in unrelated blood donors with negative Rh(D) / 实用医学杂志

Objective To investigate the distribution of weak D and Del phenotype in unrelated blood donors with negative Rh (D). Methods Four hundred and nineRhD (-) unrelated donors were screened by the saline agglutination method. Type weak D was detected by the indirect antiglobulin test, and type Del was detected by absorb radiation method. Results In 409 RhD (-) samples, 27 cases (6.61%) of type weak D were checked out, and 61 cases (14.91%) were type Del and 321 cases (78.48%) were corroborate for RhD (-), In the confirmed RhD (-) blood donors, phenotype ccee was most (49.14%), followed by Ccee (23.47%), People with phenotype ccEe accounted for 4.16%in type weak D, followed by Ccee (1.71%). Ccee accounted for 10.02%in type Del, and Ccee accounted for 1.71%. Conclusion RhD (-) donors screened by regular testing should adopt a more sensitive test for verifing type weak D or Del. In order to ensure the security of blood transfusion, people with type weak D and Del should be regarded as RhD positive blood donors, and the RhD negative people deemed to be recipients.