Faculty of Veterinary Medicine, Institute of Parasitology, Centre for Infectious Medicine, University of Leipzig, 04103Leipzig, Germany.
Department of Veterinary Medicine, College of Agriculture, Can Tho University, 900000Can Tho, Vietnam.
Parasitology. 2020 Sep;147(10):1065-1070. doi: 10.1017/S0031182020000724. Epub 2020 May 4.
The transfection of Cryptosporidium represents a major challenge, and current protocols are based on electroporation of freshly excysted sporozoites using a rather large amount of plasmid DNA which typically has a very poor yield. In this study, we report a fast and simple protocol for transfection of Cryptosporidium parvum that takes advantage of the DNA condensing power of the poly cationic polymer polyethylenimine (PEI) and the gene delivery property of the short cell-penetrating peptide octaarginine. Our novel protocol requires a very low amount of plasmid DNA and does not necessitate special laboratory equipment to be performed. Transfection appears to be more efficient in oocysts just triggered for excystation than the excysted sporozoites. Altogether, the application of octaarginine with PEI allows efficient transfection. To the best of our knowledge, this is the first report on an electroporation-free protocol for transfection of sporozoites of a Cryptosporidium species.
隐孢子虫的转染是一个主要的挑战，目前的方案是基于对刚孵出的孢子进行电穿孔，使用相当大量的质粒 DNA，其通常具有非常低的产量。在这项研究中，我们报告了一种快速而简单的隐孢子虫 parvum 转染方案，该方案利用聚阳离子聚合物聚乙烯亚胺 (PEI) 的 DNA 凝聚能力和短细胞穿透肽八聚精氨酸的基因传递特性。我们的新方案需要非常少量的质粒 DNA，并且不需要特殊的实验室设备来进行。转染似乎在刚刚被触发进行孵出的卵囊中转染效率更高，而不是孵出的孢子。总的来说，PEI 与八聚精氨酸的联合应用可实现有效的转染。据我们所知，这是第一个关于隐孢子虫种的孢子无电穿孔转染方案的报告。
