• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Harmful Algae >Sensitive and rapid detection of microcystin synthetase E Gene (mcyE) by loop-mediated sothermal amplification: A new assay for detecting the potential microcystin-producing Microcystis in the aquatic ecosystem
【24h】

Sensitive and rapid detection of microcystin synthetase E Gene (mcyE) by loop-mediated sothermal amplification: A new assay for detecting the potential microcystin-producing Microcystis in the aquatic ecosystem

机译：通过环介导的热扩增灵敏快速检测微囊藻毒素合成酶E基因（mcyE）：一种用于检测水生生态系统中潜在产生微囊藻毒素的微囊藻的新方法

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

In order to develop a new molecular technique that has the potential to assist with monitoring and management of water bodies for potential microcystin producing cyanobacterial species that occur in mixed populations in many regions of the world, we designed a new loop-mediated isothermal amplification (LAMP) assay based on microcystin biosynthesis genes. Four sets of primers were designed to recognize six distinct sequences on target the mcyE gene that encodes a protein (McyE) being responsible to catalyze the addition of D-glutamate to Adda. One set (MCYE2) was selected as the most appropriate set of primers for its rapid detection. The specificity and sensitivity of the primers in the LAMP reactions for mcyE detection were determined. Two methods, namely, monitoring of turbidity and addition of calcein to the reaction tube, were used to determine negative and positive results. The results showed that target DNA was amplified and visualized by the two detection methods within 40 min at an isothermal temperature of 61 ℃. For the sensitivity of LAMP, the detection limit was 8.5 pg/μl (approximately 17 pg) DNA. The eleven microcystin producing and four non-toxic cyanobacterial strains were selected for testing of specificity. The results of the amplification were positive with all microcystin-producing strains tested and not with four non-toxic strains, which showed that the primers had good levels of specificity. For testing the application of LAMP assay in the aquatic ecosystem, seven environmental samples from ponds and lakes in Ningbo City were also analyzed using the LAMP targeting the mcyE gene as well as an ELISA assay. Compared with these results of ELISA assay, LAMP assay is satisfied. All of these validated LAMP method being fast, simple and low in cost is a potentially valuable means for potential toxic of cyanobacterial blooms detection, especially for routine monitoring purposes in future.
机译：为了开发一种新的分子技术，该技术有可能有助于监测和管理水体中存在于世界许多地区的混合种群中的潜在微囊藻毒素生产蓝细菌物种，我们设计了一种新的环介导的等温扩增（LAMP） ）基于微囊藻毒素生物合成基因的分析。设计四组引物以识别靶标编码蛋白质（McyE）的mcyE基因上的六个不同序列，该蛋白质负责催化D-谷氨酸向Adda的添加。选择一组（MCYE2）作为最适合其快速检测的引物。确定了用于mcyE检测的LAMP反应中引物的特异性和敏感性。监测浊度和向反应管中添加钙黄绿素两种方法用于确定阴性和阳性结果。结果表明，两种检测方法均能在61℃等温下40min内扩增并可视化靶DNA。对于LAMP的灵敏度，检测限为8.5 pg /μl（约17 pg）DNA。选择了11种产生微囊藻毒素的菌株和4种无毒的蓝细菌菌株来测试特异性。在所有测试的产生微囊藻毒素的菌株中，扩增结果均为阳性，而在四种无毒菌株中则没有，这表明引物具有良好的特异性。为了测试LAMP分析在水生生态系统中的应用，还使用靶向mcyE基因的LAMP和ELISA分析了宁波市池塘和湖泊的7个环境样品。与ELISA测定的这些结果相比，LAMP测定是满意的。所有这些经过验证的快速，简单和低成本的LAMP方法，对于蓝藻水华检测的潜在毒性，尤其是将来的常规监测目的，可能是一种有价值的手段。

著录项

  • 来源
    《Harmful Algae》 |2014年第7期|8-16|共9页
  • 作者

    Peng Zhu; Bai-Feng Zhang; Jing-Hua Wu; Chen-Yang Dang; Ya-Ting Lv; Jian-Zhong Fan; Xiao-Jun Yan;

  • 作者单位

    Key Laboratory of Applied Marine Biotechnology, Ningbo University, Ningbo 315211, China,Ningbo Branch of National Engineering Research Center for Beijing Biochip Technology, Ningbo 315201, China;

    Key Laboratory of Applied Marine Biotechnology, Ningbo University, Ningbo 315211, China;

    Key Laboratory of Applied Marine Biotechnology, Ningbo University, Ningbo 315211, China;

    Key Laboratory of Applied Marine Biotechnology, Ningbo University, Ningbo 315211, China;

    Key Laboratory of Applied Marine Biotechnology, Ningbo University, Ningbo 315211, China;

    Ningbo Branch of National Engineering Research Center for Beijing Biochip Technology, Ningbo 315201, China;

    Key Laboratory of Applied Marine Biotechnology, Ningbo University, Ningbo 315211, China,Ningbo Branch of National Engineering Research Center for Beijing Biochip Technology, Ningbo 315201, China;

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

    Microcystin-producing Microcystis; LAMP assay; Rapid detection;

    机译：产生微囊藻毒素的微囊藻;LAMP分析;快速检测;

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