建立稳定的贝氏巴贝斯虫基因操作转染系统。
Establishment of a stable transfection system for genetic manipulation of Babesia gibsoni.
机构信息
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido, 080-8555, Japan.
Department of Protozoology, Institute of Tropical Medicine, Nagasaki University, Sakamoto 1-12-4, Nagasaki, 852-8523, Japan.
出版信息
Parasit Vectors. 2018 Apr 23;11(1):260. doi: 10.1186/s13071-018-2853-1.
BACKGROUND
Genetic manipulation techniques, such as transfection, have been previously reported in many protozoan parasites. In Babesia, stable transfection systems have only been established for bovine Babesia parasites. We recently reported a transient transfection system and the selection of promoter candidates for Babesia gibsoni. The establishment of a stable transfection system for B. gibsoni is considered to be urgent to improve our understanding of the basic biology of canine Babesia parasites for a better control of babesiosis.
RESULTS
GFP-expressing parasites were observed by fluorescence microscopy as early as two weeks after drug selection, and consistently expressed GFP for more than 3 months without drug pressure. Genome integration was confirmed by PCR, sequencing and Southern blot analysis.
CONCLUSIONS
We present the first successful establishment of a stable transfection system for B. gibsoni. This finding will facilitate functional analysis of Babesia genomes using genetic manipulation and will serve as a foundation for the development of tick-Babesia and host-Babesia infection models.
背景
转染等基因操作技术已在许多原生动物寄生虫中得到报道。在巴贝虫中，只有牛巴贝虫寄生虫的稳定转染系统得以建立。我们最近报道了一种暂时性转染系统和巴贝西虫候选启动子的筛选。建立稳定的 B. gibsoni 转染系统被认为是紧迫的，以改善我们对犬巴贝斯虫寄生虫基本生物学的理解，从而更好地控制巴贝斯虫病。
结果
药物选择后两周，通过荧光显微镜观察到表达 GFP 的寄生虫，并且在没有药物压力的情况下持续表达 GFP 超过 3 个月。通过 PCR、测序和 Southern blot 分析证实了基因组整合。
结论
我们首次成功建立了 B. gibsoni 的稳定转染系统。这一发现将有助于使用遗传操作对巴贝斯虫基因组进行功能分析，并为蜱-巴贝斯虫和宿主-巴贝斯虫感染模型的发展奠定基础。
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