title: Exogenous gene can be integrated into Nosema bombycis genome by mediating with a non-transposon vector.
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.
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China.|College of Bee Science, Fujian Agriculture and Forestry University, Fuzhou, 350000, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Guo
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China.|National Engineering Laboratory for Modern Silk, Soochow University, Suzhou, People's Republic of China.
authors.forename: Guangli
authors.initials: G
authors.lastname: Cao
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China.
authors.forename: Yahong
authors.initials: Y
authors.lastname: Lu
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China.|National Engineering Laboratory for Modern Silk, Soochow University, Suzhou, People's Republic of China.
authors.forename: Renyu
authors.initials: R
authors.lastname: Xue
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China.
authors.forename: Dhiraj
authors.initials: D
authors.lastname: Kumar
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China.|National Engineering Laboratory for Modern Silk, Soochow University, Suzhou, People's Republic of China.
authors.forename: Xiaolong
authors.initials: X
authors.lastname: Hu
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou, 215123, China. gongcl@suda.edu.cn.|National Engineering Laboratory for Modern Silk, Soochow University, Suzhou, People's Republic of China. gongcl@suda.edu.cn.
authors.forename: Chengliang
authors.initials: C
authors.lastname: Gong
mesh_terms: D000818:Animals; D061046:Biological Control Agents; D012831:Bombyx / Q000382:microbiology*; D002460:Cell Line; D017931:DNA Primers; D005822:Genetic Vectors / Q000235:genetics; D016681:Genome, Fungal / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics*; D016881:Microsporidiosis / Q000382:microbiology; D016818:Nosema / Q000235:genetics*; D016133:Polymerase Chain Reaction; D013172:Spores, Fungal / Q000235:genetics
chemical_list: D061046:Biological Control Agents; D017931:DNA Primers; D049452:Green Fluorescent Proteins
keywords: BmN cells; Bombyx mori; Fluorescence; Transfection; Transgenic Nosema bombycis; pIZT/V5-His vector