• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>RSC Advances >Helix loop-mediated isothermal amplification of nucleic acids
【24h】

Helix loop-mediated isothermal amplification of nucleic acids

机译：螺旋环介导的核酸等温扩增

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Isothermal nucleic acid amplification has played a key role in the point of care test (POCT). In this study, a helix loop-mediated isothermal amplification (HAMP) method with high specificity, efficiency and rapidity was developed. The MERS-Cov orf1b gene was chosen for the validation and optimization of HAMP. The HAMP analysis was performed at a constant temperature of 61-65 degrees C and yielded a self-primed spiral structure with no introduction of exogenous gene sequence by two pairs of specially designed primers. The primers for helix loop formation were composed of two complementary primers including the helix forward primer and the helix reverse primer, the 3 ends of which were complementary to their respective target nucleic acids. HAMP assay can be monitored by fluorescence signals with the addition of Eva Green in the reaction mixture. In addition, an accelerated HAMP was developed after the addition of acceleration probe, which could be finished within 75 min with a sensitivity of 10 copies per reaction. Further, a reverse transcription-HAMP (RT-HAMP) was proven to be feasible for RNA detection by combining the reverse transcriptase with DNA polymerase. Finally, both the HAMP and RT-HAMP assay were visually conducted by using Hydroxynaphthol blue (HNB) as a chromogenic indicator. All in all, it is suggested that the HAMP assay would have great potential in POCT applications.
机译：等温核酸扩增在护理时间点（POCT）发挥了关键作用。在该研究中，开发了一种具有高特异性，效率和快速性的螺旋回介质的等温扩增（HAMP）方法。选择MERS-COV ORF1B基因用于康普的验证和优化。在恒定温度为61-65℃的恒定温度下进行跳跃分析，得到自引导螺旋结构，没有引入外源基因序列的两对特别设计的引物。为螺旋环形成的引物由两个互补的引物包括螺旋正向引物和所述螺旋反向引物，所述3周'端，其中是它们各自的靶核酸互补。可以通过在反应混合物中加入EVA绿色的荧光信号来监测HAMP测定。另外，在添加加速探针后开发加速恒速，其可以在75分钟内完成，灵敏度为每次反应10份。此外，证明通过将逆转录酶与DNA聚合酶组合来证明RNA检测是可行的逆转录转录（RT-HAMP）。最后，通过使用羟基萘酚蓝（HNB）作为发色指示剂，目视进行HAMP和RT-HAMP测定。总而言之，建议汉普测定在POCT应用中具有很大的潜力。

著录项

  • 来源
    《RSC Advances》 |2018年第34期|共5页
  • 作者

    Mao Rui; Qi Lifei; Wang Zhuo; Liu Hongtao; Du Yuguang;

  • 作者单位

    Chinese Acad Sci Inst Proc Engn Key Lab Biopharmaceut Prod &

    Formulat Engn State Key Lab Biochem Engn PLA Beijing 100190 Peoples R China;

    Chinese Acad Sci Inst Proc Engn Key Lab Biopharmaceut Prod &

    Formulat Engn State Key Lab Biochem Engn PLA Beijing 100190 Peoples R China;

    Chinese Acad Sci Inst Proc Engn Key Lab Biopharmaceut Prod &

    Formulat Engn State Key Lab Biochem Engn PLA Beijing 100190 Peoples R China;

    Chinese Acad Sci Inst Proc Engn Key Lab Biopharmaceut Prod &

    Formulat Engn State Key Lab Biochem Engn PLA Beijing 100190 Peoples R China;

    Chinese Acad Sci Inst Proc Engn Key Lab Biopharmaceut Prod &

    Formulat Engn State Key Lab Biochem Engn PLA Beijing 100190 Peoples R China;

  • 收录信息
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 化学;
  • 关键词

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号