A Simple and Cost-Effective Method to Transfect Small Interfering RNAs Into Pancreatic Cancer Cell Lines Using Polyethylenimine
A Simple and Cost-Effective Method to Transfect Small Interfering RNAs Into Pancreatic Cancer Cell Lines Using Polyethylenimine
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DOI：
10.1097/mpa.0b013e3181f7e41c
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发表时间：
2011-01-01
期刊：
影响因子：
2.9
通讯作者：
Schneider, Guenter
中科院分区：
文献类型：
--
作者：
Wirth, Matthias;Fritsche, Petra;Schneider, Guenter
目的：RNA干扰是功能基因组学研究中不可或缺的工具。由于siRNA介导的RNA干扰的瞬时性质，转染试剂的连续使用是强制性的。由于转染试剂昂贵，因此必须考虑具有成本效益的替代品。在这项研究中，我们描述了一种基于聚乙烯亚胺的siRNA转染方案为胰腺癌细胞株。方法：为确定基于聚乙烯亚胺的转染效率，FAM标记的siRNA转染到几个胰腺癌细胞株，随后通过流式细胞术分析。通过蛋白质印迹在蛋白质水平上确定2种siRNA的有效敲低。结果：聚乙烯亚胺可用于没有明显的细胞形态学变化，在人类和小鼠胰腺癌细胞系的毒性是可以忽略不计的转染试剂的毒性。所用细胞系的转染效率在83%至98%之间。蛋白质水平的敲低与市售转染试剂相当。聚乙烯亚胺和siRNA的浓度，孵育时间，和细胞密度是determinates的transfection efficiency.Conclusions：聚乙烯亚胺是一个合适的和成本效益的替代用于将siRNA转染到胰腺癌细胞。
Objectives: RNA interference, an indispensable tool in functional genomics, can be induced by small interfering RNAs (siRNAs). Because of the transient nature of siRNA-mediated RNA interference, the continuous use of transfection reagents is mandatory. Because transfection reagents are expensive, cost-effective alternatives must be considered. In this study, we describe a polyethylenimine-based siRNA transfection protocol for pancreatic cancer cell lines.Methods: For determination of polyethylenimine-based transfection efficiency, a FAM-labeled siRNA was transfected into several pancreatic cancer cell lines and subsequently analyzed by flow cytometry. The effective knockdown of 2 siRNAs was determined on the protein level by Western blot. Toxicity of the transfection reagent was analyzed by viability assays.Results: Polyethylenimine can be used without overt cellular morphological changes, and toxicity is negligible in human and murine pancreatic cancer cell lines. Transfection efficiencies ranged between 83% and 98% in the cell lines used. The knockdown at the protein level was comparable to commercially available transfection reagents. Polyethylenimine and siRNA concentrations, incubation time, and cell density are determinates of the transfection efficiency.Conclusions: Polyethylenimine is a suitable and cost-effective alternative for transfecting siRNAs into pancreatic cancer cells.