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• 方法与技术 • 上一篇    

采用RAPD-PCR技术评估果蝇肠道噬菌体多样性

杜蓓蓓1,2,孙浩1,杨伟超1,董玉玲1,2,徐慧1*   

  1. 1中国科学院沈阳应用生态研究所, 沈阳 110016;2中国科学院大学, 北京 101400)
  • 出版日期:2019-11-10 发布日期:2019-11-10

Assessment of phage diversity in Drosophila melanogaster intestine by RAPD-PCR.

DU Bei-bei1,2, SUN Hao1, YANG Wei-chao1, DONG Yu-ling1,2, XU Hui1*   

  1. (1Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang 110016, China; 2University of Chinese Academy of Sciences, Beijing 101400, China).
  • Online:2019-11-10 Published:2019-11-10

摘要: 噬菌体作为微生物群落结构和功能的主要驱动力之一,研究其在动物肠道中的生态进化过程具有重要意义。本研究以黑腹果蝇为模型,探究以随机多态性DNA聚合酶链式反应(RAPD-PCR)病毒指纹技术评估肠道噬菌体的多样性。结果表明,野生型黑腹果蝇肠道噬菌体的多样性高于InR突变体黑腹果蝇。InR突变体果蝇在孵化为成虫后,可能经历一段短期的肠道噬菌体扰动,与第3天相比,第15天的肠道噬菌体多样性显著降低,而健康的野生型果蝇则没有明显变化。进一步利用落射荧光显微镜测量噬菌体和细菌的丰度,在第3天和15天,野生型果蝇游离噬菌体和细菌均处于稳定状态。第3天InR突变体果蝇肠道细菌的丰度显著降低,病毒与细菌丰度的比值(VBR)明显高于其他组;第15天InR突变型果蝇噬菌体的多样性降低,VBR明显低于各处理组。本文证实了InR突变体果蝇肠道噬菌体生态学特征与野生型存在一定的差异,也表明RAPD-PCR病毒指纹技术可以实现对肠道病毒多样性的快速评估。

关键词: 生物硝化-反硝化作用, 生物地球化学过程, 富营养化, 生态水文过程, 植物吸收

Abstract: As one of the main drivers of microbial community structure and function, virus is of great significance in studying the ecologically evolutionary processes of microbial community in animal’s intestine. In this study, the ecological characteristics of intestinal phage were explored using Drosophila melanogaster as a model with random PCR amplification of polymorphic DNA (RAPD-PCR) virus fingerprints technique. The results showed that the diversity of phage in wild-type D. melanogaster was higher than that of InR mutant D. melanogaster model. InR mutant D.melanogaster intestinal phage may undergo a short-term disturbance after hatching into adults, with the intestinal phage diversity of day 15 being significantly lower than that on day 3. In contrast, no significant changes were found in healthy wild-type fruit flies. The abundances of phages and bacteria were measured using an epifluorescence microscope. The wild-type D.melanogaster free phages and bacteria on day 3 and day 15 were all at a stable state. On the third day, the abundance of the InR mutant D.melanogaster intestinal bacteria was significantly reduced, and the virusbacteria abundance ratio (VBR) was significantly higher than other groups. On the 15th day, the diversity of InR mutant D.melanogaster phage was reduced, and VBR was also significantly lower than other groups. Therefore, the results of this study indicated that there was a significant difference between the InR mutant D.melanogaster phage and the wild-type phage. Moreover, the random polymorphic DNA polymerase chain reaction (RAPD-PCR) virus fingerprint technology is applicable for the rapid assessment of enterovirus diversity.

Key words: biological nitrification-denitrification, biogeochemical process, eutrophication, eco-hydrological process, plant assimilation.