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目的探讨建立的RhD mRNA剪接体多态性检测技术的有效性与实用性。方法采用设计的两对引物扩增全长RhD mRNA外显子,完全避开RHCE基因的干扰,特异性扩增并分析了随机选取的非血缘关系50例标本的RhDmRNA剪接情况。结果两对引物能扩增RhD mRNA剪接体的全长外显子基因并分析出RHD基因剪接体的多态性,扩增出4种剪接体的特异性:exon-1-3扩增产物:415 bp;exon-3-10扩增产物均由3种不同组合构成:产物一:约880 bp,产物二:约750 bp;产物三:约550 bp。50例标本:exon-1-3扩增产物:全部无缺失;exon-3-10扩增产物:带1无缺失,带2缺exon-7的占总标本27例、缺exon-7,8的占总标本6例、缺exon-8,9的占总标本12例、缺exon-7,9的占总标本5例,带3全部缺exon-7,8,9。结论建立的RhD mRNA剪接体多态性检测技术可以准确扩增RhD mRNA全长基因,并有效分析该基因剪接体的多态性。
Objective To investigate the validity and practicability of the established detection of RhD mRNA splicing polymorphism. Methods Two pairs of primers were designed to amplify the full-length RhD mRNA exon and completely avoid the interference of RHCE gene. RhD mRNA splicing was randomly amplified and analyzed in randomly selected non-blood relationship 50 specimens. Results Two pairs of primers could amplify the full-length exon gene of RhD mRNA splicing gene and analyze the polymorphism of RHD gene splice variant. The specificity of four splice variants was amplified: exon-1-3 amplification product: 415 bp; exon-3-10 amplification products are composed of three different combinations: product one: about 880 bp, product two: about 750 bp; product three: about 550 bp. 50 samples: exon-1-3 amplification products: all without deletion; exon-3-10 amplification products: 1 without deletion, with 2 missing exon-7 accounted for 27 cases of total specimens, the lack of exon-7,8 Of the total specimens in 6 cases, the absence of exon-8,9 accounted for 12 specimens of the total specimens, the absence of exon-7,9 accounted for 5 specimens of the total specimens, with 3 all the lack of exon-7,8,9. Conclusion The established RhD mRNA splice variant detection method can accurately amplify the full-length RhD mRNA gene and effectively analyze the polymorphism of the splice variant.