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Exogenous gene can be integrated into Nosema bombycis genome by mediating with a non-transposon vector

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Abstract

Nosema bombycis, a microsporidium, is a pathogen of pebrine disease of silkworms, and its genomic DNA sequences had been determined. Thus far, the research of gene functions of microsporidium including N. bombycis cannot be performed with gain/loss of function. In the present study, we targeted to construct transgenic N. bombycis. Therefore, hemocytes of the infected silkworm were transfected with a non-transposon vector pIZT/V5-His vector in vivo, and the blood, in which the hemocyte with green fluorescence could be observed, was added to the cultured BmN cells. Furthermore, normal BmN cells were infected with germinated N. bombycis, and the infected cells were transfected with pIZT/V5-His. Continuous fluorescence observations exposed that there were N. bombycis with green fluorescence in some N. bombycis-infected cells, and the extracted genome from the purified N. bombycis spore was used as templates. PCR amplification was carried out with a pair of primers for specifically amplifying the green fluorescence protein (GFP) gene; a specific product representing the gfp gene could be amplified. Expression of the GFP protein through Western blotting also demonstrated that the gfp gene was perfectly inserted into the genome of N. bombysis. These results illustrated that exogenous gene can be integrated into N. bombycis genome by mediating with a non-transposon vector. Our research not only offers a strategy for research on gene function of N. bombycis but also provides an important reference for constructing genetically modified microsporidium utilized for biocontrol of pests.

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Acknowledgments

We gratefully acknowledge the financial support of the National Natural Science Foundation of China (31272500), the National Basic Research Program of China (973 Program, 2012CB114600), the Specialized Research Fund for the Doctoral Program of Higher Education (20113201130002), and a project funded by the Priority Academic Program of Development of Jiangsu Higher Education Institutions. We thank Dr. Dhiraj Kumar for help with language polishing.

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Authors and Affiliations

  1. School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China

    Rui Guo, Guangli Cao, Yahong Lu, Renyu Xue, Dhiraj Kumar, Xiaolong Hu & Chengliang Gong

  2. College of Bee Science, Fujian Agriculture and Forestry University, Fuzhou, 350000, China

    Rui Guo

  3. National Engineering Laboratory for Modern Silk, Soochow University, Suzhou, People’s Republic of China

    Guangli Cao, Renyu Xue, Xiaolong Hu & Chengliang Gong

Authors
  1. Rui Guo
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  2. Guangli Cao
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  3. Yahong Lu
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  4. Renyu Xue
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  5. Dhiraj Kumar
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  6. Xiaolong Hu
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  7. Chengliang Gong
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Correspondence to Chengliang Gong.

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Rui Guo, Guangli Cao, and Yahong Lu contributed equally to this work.

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Guo, R., Cao, G., Lu, Y. et al. Exogenous gene can be integrated into Nosema bombycis genome by mediating with a non-transposon vector. Parasitol Res 115, 3093–3098 (2016). https://doi.org/10.1007/s00436-016-5064-8

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  • DOI: https://doi.org/10.1007/s00436-016-5064-8

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