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PCR在HBV核酸定量检测中的应用
  • ISSN号:1673-6273
  • 期刊名称:《现代生物医学进展》
  • 时间:0
  • 分类:R512.62[医药卫生—临床医学;医药卫生—内科学]
  • 作者机构:[1]第四军医大学唐都医院病理科第四军医大学唐都医院妇产科,陕西西安710038
  • 相关基金:国家自然科学基金项目(30672013;81372226);陕西省科技统筹创新工程(2011KTCL03-11)
中文摘要:

目的:探索一种简便定量分析系统,通过检测HBV携带者血清中的HBV X区DNA,fRNA和trRNA拷贝数,为提高隐匿性感染期低复制状态HBV检测效果提供可能。方法:从血清中提取HBV DNA和RNA,对Core区、X区DNA进行PCR扩增,使用半巢式PCR法对fRNA、trRNA在同一离心管中进行反转录并扩增,选取大小相近的质粒作为竞争模板对其进行定量。并对拉米夫定干预治疗14周前后的患者血清中HBV XDNA、Core DNA、fRNA和trRNA拷贝数进行检测与比较。所有检测结果均通过southern杂交进行验证。结果:建立了针对DNA的定量分析系统及针对RNA的RT-PCR定量分析系统,并且阐明了阿米夫定治疗前后HBV DNA和X区RNA结构和数量的变化。治疗前治疗后DNA和RNA的拷贝数均下降。Core DNA下降显著,为103-104倍,而XDNA拷贝数下降102倍。而fRNA和trRNA仅有小幅下降,为10倍左右。结论:可以通过竞争性PCR方法对血清中HBV DNA和RNA进行定量检测,以期为HBV病毒的诊断提供更充足依据。

英文摘要:

Objective: To build a feasible quantitative analysis system to improve the possibility of detection of the HBV at the occult infection according to the copy numbers of HBV XDNA, fRNA and trRNA in the serum. Methods: HBV DNA and RNA were extracted from serum samples. HBV Core DNA, X DNA were amplified by polymerase chain reaction (PCR), and the heminested PCR were used in the reverse transcriptase-polymerase chain reaction (RT-PCR) system for fRNA and trRNA. They were marked by the plasmid sized around them as competitive templates. The copy numbers of HBV XDNA, Core DNA fRNA and trRNA in the serum treated by lamivudine for 14 weeks were compared with before. Results were verified by the southern hybridization. Results: The DNA quantitative system and RNA RT-PCR quantitative system were established. The exchange of the HBV DNA and RNA in number and structure were clarified. The copy number of DNA and RNA decreased after lamivudine treatment. Core DNA decreased dramatically by 10^3-10^4 times, and X DNA decreased 10^2 times. While fRNA and trRNA decreased around 10 times. Conclution: HBV DNA and RNA can be detected by the competitive PCR and it is likely to be useful to the RBV virus examination.

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期刊信息
  • 《现代生物医学进展》
  • 中国科技核心期刊
  • 主管单位:黑龙江省卫生厅
  • 主办单位:黑龙江省红十字医院 黑黑龙江省红十字医院 黑龙江省森林工总医院
  • 主编:申宝忠
  • 地址:哈尔滨市南岗区花园街184号403
  • 邮编:150001
  • 邮箱:biomed_54@126.com
  • 电话:0451-82583800 53658268
  • 国际标准刊号:ISSN:1673-6273
  • 国内统一刊号:ISSN:23-1544/R
  • 邮发代号:14-12
  • 获奖情况:
  • 国内外数据库收录:
  • 被引量:33230