引用本文:单雪峰,谢素兰,黄爱龙,胡 源.应用肽核酸反向杂交技术检测HBV阿德福韦耐药突变株[J].重庆医科大学学报,2014,38(7):943~946
应用肽核酸反向杂交技术检测HBV阿德福韦耐药突变株
Application of peptide nucleic acid reverse hybridization technology in detecting adefovir-resistant mutations in hepatitis B virus
DOI:
中文关键词:  肽核酸  反向杂交  乙型肝炎病毒  阿德福韦酯
英文关键词:peptide nucleic acid  reverse hybridization  hepatitis B virus  adefovir dipivoxil
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作者单位
单雪峰,谢素兰,黄爱龙,胡 源 重庆医科大学感染性疾病分子生物学教育部重点实验室重庆 400016 
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中文摘要:
      目的:建立一种肽核酸(peptide nucleic acid,PNA) 反向杂交技术,用于检测乙肝病毒阿德福韦酯(adefovir dipivoxil,ADV)常见耐药突变位点。方法:利用反向杂交技术,设计出特异性的PNA探针,与PCR扩增后带有生物素标记的样本进行杂交;通过对探针浓度,杂交温度和时间等条件的优化建立起最佳反应条件。应用该技术检测了临床ADV治疗慢性乙肝患者的72例血清样本,与测序法比较了其检测的准确性。结果:(1)通过共价结合能够将PNA探针固定在尼龙膜上,特异性检测ADV常见耐药突变位点。(2)与直接测序法相比,PNA反向杂交技术检测准确性为97.88%。结论:通过优化杂交反应,建立起快速、灵敏的检测乙肝病毒ADV常见耐药突变位点的PNA反向杂交技术。
英文摘要:
      Objective:To detect adefovir-resistant mutants of hepatitis B virus(HBV) using peptide nucleic acid(PNA) probes. Methods:Specific PNA probes were designed for genotypes B,C D,which were then hybrized with biotinylated PCR products. Different hy-bridization factors such as probe concentration,hybridization temperature and time were investigated to establish the optimization conditions. Finally,72 hepatitis B virus(HBV) clinical samples with adefovir dipivoxil treatment were investigated using PNA hybriza-tion assay. Its usefulness was validated by comparing with sequencing data. Results:(1)PNA probes could be fixed on the membrane by cross link and used for reverse hybridization. (2)Complete concordance of PNA hybrization assays was 97.88% with direct sequenc-ing. Conclusion:We develop an assay using PNA probes to detect adefovir-resistant mutants of HBV.
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