Transfectamine mRNA转染试剂

样品实验方案

简要概述

1.准备转染细胞
2.制备Transfectamine mRNA转染试剂-RNA 混合物
3.将Transfectamine mRNA转染试剂-RNA 混合物添加到细胞培养物中
4.过夜培养细胞
5.用合适的方法分析转染效率

 

细胞准备

1.在转染时将细胞培养至 ~ 90% 汇合。
2.转染前更换新鲜培养基。例如，6 孔板每孔更换 2 mL 培养基，10 cm 板更换 6 mL 培养基。

 

工作溶液配制

1.Transfectamine mRNA转染试剂-RNA 混合物
2.将 2.5 µg mRNA 与 200 µL 无血清培养基混合。
3.在步骤 1 中加入 7.5 µL Transfectamine mRNA转染试剂。
4.混匀并在室温下孵育 20 分钟。
注意：Transfectamine mRNA转染试剂与 mRNA 的比例需要针对不同的细胞系进行优化。 通常，Transfectamine mRNA转染试剂 (µL) 与 mRNA (µg) 的比率 =（3 至 5 µL）至 1 µg。

6孔板样品方案

组分	6孔板（每孔）
新鲜培养基	2ml
纯化的mRNA	-2.5ug
无血清培养基	200ul
Transfectamine mRNA转染试剂	-7.5ul

 

操作步骤

1.转染方案

1.1将Transfectamine mRNA转染试剂 - mRNA 混合物加入培养板并培养过夜。
注意：重组蛋白表达可以在转染后的8小时内检测到。 转染后约24小时可观察到表达水平。

图 1. HeLa 细胞中的转染效率比较（上图）和细胞毒性比较（下图）。 HeLa 细胞在 6 孔板中培养至~90% 汇合。 2.5 µg mRNA 分别用 Lipofactamin MessengerMAX 和Transfectamine mRNA转染试剂转染。 转染后 18 小时，使用带有 FITC 通道的荧光显微镜（上图）拍摄图像。 Lipofactamin MessengerMAX 和Transfectamine mRNA转染试剂的转染效率相似。用Transfectamine mRNA转染试剂转染的细胞看起来比用 Lipofatamin MessengerMAX 转染的细胞健康得多（下图）。

 

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