摘要
目的通过检测慢性乙型肝炎重型(chronic severe hepatitis B,CSHB)患者趋化因子CXCL10[干扰素γ诱导蛋白(IFN γ-inducible protein,IP-10)]基因启动子区G-201A位点的变异率,探讨PCR-限制性片段长度多态性(restriction fragment length polymorphism,RFLP)法用于批量单核苷酸多态性(single nucleotide polymorphism,SNP)分型鉴定的应用价值。方法选取解放军第三〇二医院的200例CSHB患者(CSHB组)为研究对象,300例健康体检者作为正常对照(normal control,NC)组。收集患者EDTA-K2抗凝全血,提取白细胞DNA,采用PCR-RFLP法进行检测,对其中185例[CSHB组93例(46.5%),NC组92例(30.7%)]样本(占总样品的37%)用DNA测序法进行验证。结果 185例样本用DNA测序法进行验证,经Kappa检验k=0.937,P<0.05,说明二种检测结果之间的一致性具有统计学意义,二种方法的一致率为97.84%,符合体外诊断试剂临床试验指导方案中关于二种方法一致性的专业要求(一致率≥95%)。CSHB组趋化因子IP-10相关的G-201A的突变率为21.28%、NC组为13.82%,差异有统计学意义(P<0.05)。结论 CSHB组患者的趋化因子IP-10基因启动子区G-201A位点的突变率明显高于NC组。本实验结果表明,PCR-RFLP法经过直接测序法验证结果可靠,可以用于标本的SNP分型。
Objective To investigate the application of restriction fragment length polymorphism （RFLP） method to the identi- fication of the volume single nucleotide polymorphism （SNP） genotyping by detecting the mutation rate of chemokine （C-X-C motif） ligand 10 （CXCLIO）, also named as IFN y-inducible protein （IP-10）, gene promoter G-201A site in patients with chronic severe hepatitis B （CSHB）. Methods Totally 200 CSHB patients in 302 hospital of PLA （CSHB group） and 300 normal controls （NC group） were included in the study. The whole blood anticoagulated with EDTA-K2 was collected, leukocyte DNA was extracted, and PCR-RFLP method was used to detect polymorphisms. Meanwhile, 185 samples including 93 （46.5%） from CSHB group and 92 （30.7%） from NC group were verified by the method of DNA sequencing. Results The agreement between PCR-RFLP method and direct sequencing analysis was statistically significant （k =0.937, P〈O.05）. The agreement rate of the two methods （97.84%） was in accordance with professional requirements （≥95%）. The mutation rates of G-201A related to chemokine IP-10 was 21.28% in CSHB group and 13.82% in NC group, and the difference between them was significant （P〈O.05）. Conclusions The mutation rate of chemokine IP-IO gene promoter G-201A site in CSHB group is higher than that in NC group. Our study shows that PCR-RFLP method verified by direct sequencing analysis is reliable, and can be used for the volume SNP genotyping identification.
出处
《传染病信息》
2011年第2期100-102,共3页
Infectious Disease Information
基金
国家"十一五"科技重大专项(2008ZX10002-005-6)
国家重点基础研究发展计划课题(2007CB512803)
关键词
趋化因子类
肝炎
乙型
慢性
多态现象
单核苷酸
多态现象
限制性片段长度
chemotactic factors
hepatitis B, chronic
polymorphism, single nucleotide
polymorphism, restriction fragmentlength
