Polyethyleneimine (PEI) Mediated siRNA Gene Silencing in the Schistosoma mansoni Snail Host, Biomphalaria glabrata
Polyethyleneimine (PEI) Mediated siRNA Gene Silencing in the Schistosoma mansoni Snail Host, Biomphalaria glabrata
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DOI：
10.1371/journal.pntd.0001212
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发表时间：
2011-07-01
影响因子：
3.8
通讯作者：
Ittiprasert, Wannaporn
中科院分区：
文献类型：
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作者：
Knight, Matty;Miller, Andre;Ittiprasert, Wannaporn
利用阳离子聚合物聚乙烯亚胺（PEI）介导的长双链（ds）和小干扰（si）RNA的递送，已经开发了一种在蜗牛中通过RNA干扰（RNAi）进行基因沉默的体内非侵入性技术。通过使用“模拟”荧光siRNA评估和优化细胞递送。随后，我们使用该方法用该转录物的相应siRNA或dsRNA抑制组织蛋白酶B（CathB）的表达。此外，敲低过氧化物氧还蛋白（Prx）在RNA和蛋白质水平实现与PEI介导的浸泡方法。B。光滑蜗牛是传播寄生性双殖吸虫（digenean platyhelminth）、曼氏血吸虫（Schistosoma mansoni）的重要蜗牛宿主，曼氏血吸虫在新热带地区引起血吸虫病。目前正在取得进展，以实现蜗牛的基因组序列，并揭示基因表达谱和细胞途径，使蜗牛能够预防或维持感染。使用PEI复合物，已经开发了一种方便的浸泡方法，使得能够使用dsRNA或siRNA进行功能基因敲除研究。开发的协议提供了第一个完整的生物体的宿主-寄生虫的基因功能研究所需的确定寄生虫在蜗牛宿主，这最终需要作为点破坏这种寄生虫介导的疾病的发展的关键机制。
An in vivo, non-invasive technique for gene silencing by RNA interference (RNAi) in the snail, Biomphalaria glabrata, has been developed using cationic polymer polyethyleneimine (PEI) mediated delivery of long double-stranded (ds) and small interfering (si) RNA. Cellular delivery was evaluated and optimized by using a 'mock' fluorescent siRNA. Subsequently, we used the method to suppress expression of Cathepsin B (CathB) with either the corresponding siRNA or dsRNA of this transcript. In addition, the knockdown of peroxiredoxin (Prx) at both RNA and protein levels was achieved with the PEI-mediated soaking method. B. glabrata is an important snail host for the transmission of the parasitic digenean platyhelminth, Schistosoma mansoni that causes schistosomiasis in the neotropics. Progress is being made to realize the genome sequence of the snail and to uncover gene expression profiles and cellular pathways that enable the snail to either prevent or sustain an infection. Using PEI complexes, a convenient soaking method has been developed, enabling functional gene knockdown studies with either dsRNA or siRNA. The protocol developed offers a first whole organism method for host-parasite gene function studies needed to identify key mechanisms required for parasite development in the snail host, which ultimately are needed as points for disrupting this parasite mediated disease.