摘要
目的建立一种可靠、敏感、全面的ERCC1 Asn118Asn基因多态性检测新方法,用于预测肿瘤患者对铂类的耐药性。方法以ERCC1基因第4个外显子Asn118Asn的一对引物行PCR扩增227例临床样本DNA,将荧光偏振检测技术与模板指导的末端延伸反应(TD I-FP)结合,应用探针杂交延伸反应对临床样本扩增产物进行ERCC1 Asn118Asn基因多态性检测,并以DNA序列测定验证检测结果。结果227例临床样本DNA中,119例(52.4%)ERCC1 Asn118Asn基因为C/C纯合子(AAC)基因型,88例(38.8%)为C/T杂合子基因型,20例(8.8%)为T/T纯合子(AAT)基因型。但DNA序列测定法检出杂合子基因型仅33例。结论本研究初步建立了特异性较好、操作简便、无需标记探针的临床标本ERCC1 Asn118Asn基因多态性检测新方法,有望用于肿瘤患者铂类耐药性的检测。
Objective To develop a simple and accurate method for detecting polymorphism of the excision repair cross-complementing group 1（ERCC1） Asn118Asn by template direct dye-terminator incorporation with fluorescence-polarization（TDI-FP） assay.Methods The primers of ERCC1 exon 4 were used to amplify DNA of 227 samples and the PCR products were detected by TDI-FP.A fluorescence labeled ddNTP was directly added to the probe under the direction of template,the specific PCR products.The results were measured by a fluorescence polarization reader.Results One hundred and nineteen（52.4%） patients were homozygous for C/C genotype,20（8.8%） were homozygous for T/T genotype,and 88（38.8%） were heterozygous for C/T genotype.However,only 33 patients were identified as heterozygous（C/T genotype） by DNA sequencing.The method developed in this study was more sensitive for detection of the polymorphism than DNA sequencing.Conclusions The assay developed in the present study allows a simple and accurate detection of the polymorphism of ERCC1 Asn118Asn without any use of label probes.
出处
《临床检验杂志》
CAS
CSCD
北大核心
2010年第3期197-199,共3页
Chinese Journal of Clinical Laboratory Science
基金
国家高技术研究和发展计划(2008AA02Z444)
