| 李 彤#, 蒋月丽#, 连红梅, 武予清*, 苗 进, 巩中军, 段 云.蚜虫共生细菌多重PCR检测体系的建立[J].植物保护,2020,46(2):158-163. |
| 蚜虫共生细菌多重PCR检测体系的建立 |
| Establishment of multiplex PCR detection system for symbiotic bacteria of aphids |
| 投稿时间：2019-02-21 修订日期：2019-05-09 |
| DOI：10.16688/j.zwbh.2019069 |
| 中文关键词: 沃尔巴克氏菌 杀雄菌属共生菌 蚜虫U型共生菌 多重PCR |
| 英文关键词:Wolbachia pipientis Arsenophonus Regiella insecticola multiplex PCR |
| 基金项目:国家自然科学基金（31601897）;河南省农业科学院杰出青年基金 |
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| 中文摘要: |
| 蚜虫中具有多种共生菌,使用常规PCR对它们进行检测,耗时耗力,而多重PCR可以更加高效地进行多种细菌的检测。沃尔巴克氏菌Wolbachia pipientis、杀雄菌属共生菌Arsenophonus和蚜虫U型共生菌Regiella insecticola是蚜虫中常见的3种共生菌。本研究针对沃尔巴克氏菌、杀雄菌属共生菌和蚜虫U型共生菌,分别选择以wsp基因、yaeT基因和gltA基因作为靶标,进行了多重PCR引物的设计和扩增体系的优化。结果显示,本研究建立的多重PCR体系在检测3种蚜虫常见共生菌时,具有较高的扩增特异性、准确性和直观性及较高的检测灵敏度,共生菌的最低检测浓度为104拷贝/μL,远低于共生菌在蚜虫1龄若虫总DNA中的浓度(108拷贝/μL),可以完全满足蚜虫共生菌检测工作的需要。 |
| 英文摘要: |
| Aphids harbor many bacterial symbionts. Although conventional PCR is universally used in the detection of aphid bacterial symbionts, however, it is a time-consuming way when multiple aphid bacterial symbionts should be detected in the same sample. Previous researches revealed that the multiplex PCR had high efficiency in the multiple bacteria detections. Wolbachia, Arsenophonus and Regiella insecticola are the three universal bacterial symbionts in aphids. In this study, we employ three protein coding genes, wsp, yaeT and gltA, as targets to design the multiplex PCR primers and establish an optimized detection system. The results showed that the multiplex PCR system established in this study had high amplification specificity and accuracy in the detection of the three aphid bacterial symbionts. Moreover, the minimum detection concentration of symbiotic bacteria was 104 copies/μL, lower than that of that in the total DNA of the first instar aphid, 108 copies/μL, indicating that the multiplex PCR system also had the high sensitivity in the bacterial symbionts detection, which will fully meet the requirements in the detection of aphid bacterial symbionts. |
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