一例ABO疑难血型基因分析
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摘要
目的对一例血型血清学鉴定困难的患者进行相关基因分析,鉴定血型并分析引起血型鉴定困难的原因。方法采用微柱凝胶法和试管法进行血清学血型鉴定、抗球蛋白试验和抗体筛查,PCR扩增患者ABO基因第6、7外显子区及其侧翼序列并进行Sanger测序、克隆测序,寻找突变位点进行血型鉴定。结果患者血清学血型鉴定正反结果不符,其红细胞与单克隆抗A、B、H抗体反应均呈阴性(-);血清与A、B、O型红细胞均无凝集;患者直接及间接抗球蛋白试验均为阴性;血清抗体筛查为阴性。测序发现,患者ABO血型相关基因存在多个变异位点。其中第6外显子区存在c.261delG,第7外显子区存在c.467C>CT、c.526C>CG、c.657>CT、c.703G>GA、c.796C>CA、c.803G>G。以A101基因作为标准序列进行比较,对照Blood Group Antigen Gene Mutation Database进行ABO等位基因突变分析,判断患者血型为A102/B101。结论患者红细胞上抗原减弱是血型鉴定正反不符的主要原因,DNA基因测序第6、7外显子七个点突变导致了红细胞上抗原减弱。
Objective:To investigate the gene identification of a patient with serologic blood group discrepancy,and analyze the reason and variations. Methods:Microcolumn agglutination and routine tube method were used for serologic blood group identification,and Coombs' and antibody screening were performed. Exon 6,7 and their adjacent intron region of ABO gene were amplified by polymerase chain reaction(PCR),and the PCR products were directly sequenced to identify the gene mutation.Results:The forward and reverse tests of serologic blood group identification were not consistent. The red blood cells of the patient showed negative(-)with monoclonal anti-A anti-B and anti-H. The patient's serum showed weak agglutination with type A1 red blood cells and B blood cells and type O red blood cells.The direct and indirect Coombs and antibody screening were all negative.There were heterozygous variations in exon 6(261 del)and 6 heterozygous variations in exon 7(c.467 C>CT、c.526 C>CG、c.657>CT、c.703 G>GA、c.796 C>CA、c.803 G>G)of ABO gene in the patient compared with the reference sequence of A101 allele.Based on the Blood Group Antigen Gene Mutation Database,A102/B101 subgroup alleles were identified. Conclusions:The weak of AB antigen are the main reasons of the discrepancy in blood group identification for the patient.The seven heterozygous in exons 6 and 7 of DNA gene led to AB antigen weaked.
Objective:To investigate the gene identification of a patient with serologic blood group discrepancy,and analyze the reason and variations. Methods:Microcolumn agglutination and routine tube method were used for serologic blood group identification,and Coombs' and antibody screening were performed. Exon 6,7 and their adjacent intron region of ABO gene were amplified by polymerase chain reaction(PCR),and the PCR products were directly sequenced to identify the gene mutation.Results:The forward and reverse tests of serologic blood group identification were not consistent. The red blood cells of the patient showed negative(-)with monoclonal anti-A anti-B and anti-H. The patient's serum showed weak agglutination with type A1 red blood cells and B blood cells and type O red blood cells.The direct and indirect Coombs and antibody screening were all negative.There were heterozygous variations in exon 6(261 del)and 6 heterozygous variations in exon 7(c.467 C>CT、c.526 C>CG、c.657>CT、c.703 G>GA、c.796 C>CA、c.803 G>G)of ABO gene in the patient compared with the reference sequence of A101 allele.Based on the Blood Group Antigen Gene Mutation Database,A102/B101 subgroup alleles were identified. Conclusions:The weak of AB antigen are the main reasons of the discrepancy in blood group identification for the patient.The seven heterozygous in exons 6 and 7 of DNA gene led to AB antigen weaked.
引文
[1]叶应妩,王毓三,申子瑜.全国临床检验操作规程[M].第3版.南京:东南大学出版社,2006.246-258.
[2]Yamamoto F,McNeil PD,Hzkomri S. Genomic organization of human histo-blood group ABO genes[J]. Glycobiology,1995,5:51-58.
[3]Yamamoto F. Marken J,Tsuji T,et al. cloning and characterization of DNA complementary to human UDP-GaINAc:Fuc alpha1-2 Gal alpha1-3 Gal NAc mRNA[J].J Bio Chem,1990,256:1146-1151.
[4]李慧敏,林隽峰,张帆,等.一例罕见B(A)血型的基因鉴定[J].检验医学,2014,9(9):945-947.
[5]闫有敏.凝聚胺法和微柱凝胶抗人球蛋白法在配血中的应用比较[J].实用医技杂志,2013,20(40):418-418.
[6]孟庆林,王雷萍,翁彬,等.类孟买型个体FUT1、FUT2基因测序研究[J].中国热带医学,2009,9(8):1413-1414.
[7]陈忠,徐秀玉,梁建英.白血病引起ABO血型抗原减弱若干问题探讨[J].中华血液学杂志,1986,7(8):457.
[2]Yamamoto F,McNeil PD,Hzkomri S. Genomic organization of human histo-blood group ABO genes[J]. Glycobiology,1995,5:51-58.
[3]Yamamoto F. Marken J,Tsuji T,et al. cloning and characterization of DNA complementary to human UDP-GaINAc:Fuc alpha1-2 Gal alpha1-3 Gal NAc mRNA[J].J Bio Chem,1990,256:1146-1151.
[4]李慧敏,林隽峰,张帆,等.一例罕见B(A)血型的基因鉴定[J].检验医学,2014,9(9):945-947.
[5]闫有敏.凝聚胺法和微柱凝胶抗人球蛋白法在配血中的应用比较[J].实用医技杂志,2013,20(40):418-418.
[6]孟庆林,王雷萍,翁彬,等.类孟买型个体FUT1、FUT2基因测序研究[J].中国热带医学,2009,9(8):1413-1414.
[7]陈忠,徐秀玉,梁建英.白血病引起ABO血型抗原减弱若干问题探讨[J].中华血液学杂志,1986,7(8):457.