Wang Liu, Wang Rui, Yu Yonghua, Zhang Fang, Wang Xiaofu, Ying Yibin, Wu Jian, Xu Junfeng
College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou, 310058, China.
Zhejiang Academy of Agricultural Science, Hangzhou, 310021, China.
Anal Bioanal Chem. 2016 Jan;408(2):657-62. doi: 10.1007/s00216-015-9128-x. Epub 2015 Oct 29.
The requirement of power-dependent instruments or excessive operation time usually restricts current nucleic acid amplification methods from being used for detection of transgenic crops in the field. In this paper, an easy and rapid detection method which requires no electricity supply has been developed. The time-consuming process of nucleic acid purification is omitted in this method. DNA solution obtained from leaves with 0.5 M sodium hydroxide (NaOH) can be used for loop-mediated isothermal amplification (LAMP) only after simple dilution. Traditional instruments like a polymerase chain reaction (PCR) amplifier and water bath used for DNA amplification are abandoned. Three kinds of dewar flasks were tested and it turned out that the common dewar flask was the best. Combined with visual detection of LAMP amplicons by phosphate (Pi)-induced coloration reaction, the whole process of detection of transgenic crops via genetically pure material (leaf material of one plant) could be accomplished within 30 min. The feasibility of this method was also verified by analysis of practical samples.
依赖电力的仪器要求或过长的操作时间通常限制了当前核酸扩增方法在田间检测转基因作物中的应用。本文开发了一种无需供电的简便快速检测方法。该方法省略了耗时的核酸纯化过程。用0.5M氢氧化钠(NaOH)从叶片中获得的DNA溶液只需简单稀释即可用于环介导等温扩增(LAMP)。摒弃了用于DNA扩增的传统仪器,如聚合酶链反应(PCR)扩增仪和水浴锅。测试了三种杜瓦瓶,结果表明普通杜瓦瓶效果最佳。结合通过磷酸盐(Pi)诱导显色反应对LAMP扩增产物进行可视化检测,通过遗传纯合材料(单株植物的叶片材料)检测转基因作物的整个过程可在30分钟内完成。通过实际样品分析也验证了该方法的可行性。