RHCE基因编码区全长测序方法的建立  

Establishing a sequencing method of RHCE gene coding region

作　　者：易品[1] 吴大洲[2] 彭进 伍昌林[4] 党鑫堂[4] 李剑平[5] 张印则[6] 徐华[2] 黎诚耀[1] 邵超鹏[1,4] 

机构地区：[1]南方医科大学生物技术学院输血医学系,广东广州510515 [2]陕西省血液中心 [3]西安市卫生学校 [4]深圳大学附属第一医院 [5]辽宁省血液中心 [6]深圳市血液中心

出　　处：《中国输血杂志》2015年第7期769-772,共4页Chinese Journal of Blood Transfusion

基　　金：深圳市科技创新计划(JCYJ2014041470821221JCYJ20130401110246572);深圳市第二人民医院院属重点实验室项目资助

摘　　要：目的建立简易的RHCE基因编码区全长测序方法。方法依据RHCE基因序列的特异性,设计10对特异性引物分别扩增RHCE基因的10个外显子,同时再设计10条测序引物用于测序。将该方法用于检测3例Rh阳性标本、1例Rh阴性标本、1例D--表型标本及1例弱E表型标本。结果 10对特异性引物在统一的PCR条件下成功扩增出RHCE基因的10个外显子,3例Rh阳性标本及1例Rh阴性标本的测序序列与RHCE基因标准序列一致;D--标本PCR扩增和测序结果显示缺失第3、4和第5外显子,提示该个体携带一种新的由RHCE/RHD基因交换形成的等位基因RHCE-D(3-5)-CE;弱E标本测序结果发现在第5外显子存在一处碱基突变(762G>C),其他外显子测序结果未发现变异。结论 RHCE基因编码区全长测序方法可用于一般实验室进行RHCE基因测序。Objective To establish a sequencing method of RHCE gene coding region. Methods Based on the specificity of RHCE gene,ten pairs of PCR primers and ten sequencing primers were designed. Ten pairs of PCR primers were used to amplify RHCE gene’s 10 exons. Then,ten sequencing primers were used for sequencing. Three Rh D positive samples,one Rh D negative sample,Rh deletion（ D--） sample and 1 weak E antigen sample were determined using this genotyping method. Results RHCE gene’s 10 exons can be amplified specifically under a unified PCR condition. Sequencing results of Rh D positive and 1 Rh D negative samples were consistent with standard RHCE gene sequence. In addition,a new RHCE allele RHCE-D（ 3-5）-CE was identified in D--sample and a new mutation（ 762 G 〉 C） was identified in weak E antigen sample. Conclusion RHCE gene coding region sequencing method can be used for RHCE gene sequencing analysis in general laboratory.

关 键 词：RHCE基因 序列分析 PCR D-- 弱E 

分 类 号：R457.11[医药卫生—治疗学]