[等位基因特异性引物聚合酶链反应序列分型的建立及HLA-DRB1第3外显子多态性研究]

[Establishment of allele specific primer polymerase chain reaction sequence-based typing and investigation of the polymorphism in exon 3 of HLA-DRB1].

作者信息

Zhu Fa-ming, He Yan-min, Tao Su-dan, Zhang Wei, Wang Wei, He Jun-jun, Lv Hang-jun, Yan Li-xing

机构信息

Blood Center of Zhejiang Province, Key Laboratory of Blood Safety Research of Ministry of Health of China, Hangzhou, Zhejiang, P.R. China.

出版信息

Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2010 Dec;27(6):639-43. doi: 10.3760/cma.j.issn.1003-9406.2010.06.008.

DOI:10.3760/cma.j.issn.1003-9406.2010.06.008

PMID:21154323

Abstract

OBJECTIVE

To establish the allele specific primer polymerase chain reaction sequence-based typing (PCR-SBT) and investigate the polymorphism of exon 3 of human leukocyte antigen( HLA)-DRB1.

METHODS

The fragment containing exons 2 and 3 of HLA-DRB1 gene was amplified by group specific primers. The amplified products were digested by restriction enzymes and directly sequenced in both directions. The genotype was assigned by using Assign 3.5 SBT software.

RESULTS

The exon 3 sequences of HLA-DRB108:09 and HLA-DRB112:02:01 were identified for the first time. There were 27 polymorphic sites in exon 3 among the twenty-five HLA-DRB1 alleles, which was 9.56% of all nucleotides of exon 3. The method could discriminate the HLA-DRB114:01:01/14:54 ambiguous samples, and the HLA-DRB114:01:01 was identified in the Chinese population.

CONCLUSION

The PCR-SBT method for exon 3 of HLA-DRB1 from the present study was reliable and there were polymorphisms in exon 3 of HLA-DRB1.

摘要

目的

建立等位基因特异性引物聚合酶链反应序列分型（PCR-SBT）方法并研究人类白细胞抗原（HLA）-DRB1第3外显子的多态性。

方法

采用组特异性引物扩增包含HLA-DRB1基因第2和第3外显子的片段。扩增产物用限制性内切酶消化并双向直接测序。使用Assign 3.5 SBT软件确定基因型。

结果

首次鉴定出HLA-DRB108:09和HLA-DRB112:02:01的第3外显子序列。25个HLA-DRB1等位基因的第3外显子中有27个多态性位点，占第3外显子所有核苷酸的9.56%。该方法可区分HLA-DRB114:01:01/14:54模糊样本，并在中国人群中鉴定出HLA-DRB114:01:01。