摘要
参照HLAⅡ型基因序列设计8条序列特异性引物，应用分别代表HLA－DR1－18血清学特异性的11株DNA标准品建立序列特异性PCR（PCR-SSP）和套式PCR方法，并对64例IDDM病人和72例健康对照者进行HLA－DRB1基因分型。结果：（1）方法学鉴定示各序列特异性引物均能从11株DNA标准品中扩增出相应的等位基因，并能检出来合子。除HLA－DR3组特异引物对DR1发生交叉外，其余序列特异性引物扩增中均未出现交叉现象，且重复性好；（2）样本测定显示IDDM组与对照组HLA－DR1、DR2、DR4、DR7、DR8和DR10各等位基因频率无显著差异。结果表明，PCR-SSP不仅具有较好的敏感性、特异性和重复性，而且显得简便、快速经济，有较好的应用前景。
PCR - SSP(PCR -sequence-specific primers) and nested PCR were used, and eigh sequence - specificprimers (SSP) of HLA - DR1, DR2, DR4, DR7, DR9, DR10, DR3 group (including DR3, DR8, DR1l ~ 14) alleles wer de-signed accoording to HLA class Ⅱ sequences. PCR - SSP was established with 11 DNA standards representign HLA - DRI ~ 18serological speeificiy respectively. Results showed that each of alleles of 11 DNA standaIds was successop amoped with corm-spowhng SSP. The amopcation Pattern of each SSP to 11 DNA standwh showh no cross - reaction, except HLA - DR3 group-specific primers which not only amplified the DR3, DR8,DR11 ~ 14 alleles but also amplified DR1 alleses. It is showed thatPCR - SSP is a promising HLA genotyping technique because of its high sensitivity and specificity, good reproducibility, sim-plicity, rapidity and inexpensiveness. Rational design of SSP and optimization of experiment conditions are necessary for success-ful PCR - SSP. DNA concentration in the second round amplifications is very important when nested PCR was adopted.
出处
《广东医学》
CAS
CSCD
1998年第4期245-248,共4页
Guangdong Medical Journal
基金
广东省科委重点科技攻关项目
