He Zhengmi, Su Yuting, Li Sainan, Long Pan, Zhang Ping, Chen Zuohong
College of Life Sciences, Hunan Normal University, Changsha, China.
Front Microbiol. 2019 Jul 3;10:1523. doi: 10.3389/fmicb.2019.01523. eCollection 2019.
In the present work, loop-mediated isothermal amplification (LAMP) and hyperbranched rolling circle amplification (HRCA) methods were developed to detect and distinguish different lethal species. Specific LAMP primers and HRCA padlock probes for species-specific identification and a set of universal LAMP primers for lethal species were designed and tested. The results indicated that the LAMP-based assay was able to discriminate introclade lethal species but was not able to discriminate intraclade species perfectly, while the HRCA-based assay could discriminate whether introclade or intraclade species. The universal LAMP primers were positive for 10 lethal species of section and negative for 16 species of outside section . The detection limits of LMAP and HRCA were 10 and 1 pg of genomic DNA per reaction, respectively. In conclusion, the two methods could be rapid, specific, sensitive and low-cost tools for the identification of lethal species.
在本研究中，开发了环介导等温扩增（LAMP）和超分支滚环扩增（HRCA）方法来检测和区分不同的致死物种。设计并测试了用于物种特异性鉴定的特异性LAMP引物和HRCA锁式探针以及一组用于致死物种的通用LAMP引物。结果表明，基于LAMP的检测方法能够区分进化枝内的致死物种，但不能完美区分进化枝内的物种，而基于HRCA的检测方法可以区分进化枝内或进化枝间的物种。通用LAMP引物对该组的10种致死物种呈阳性，对该组外的16种物种呈阴性。LMAP和HRCA的检测限分别为每个反应10 pg和1 pg基因组DNA。总之，这两种方法可以成为用于鉴定致死物种的快速、特异、灵敏且低成本的工具。
