生物秀首页 新闻前沿 产业资讯 实验技术 仪器教程 生物考研 资源下载 生物人 物秀商城 生物论坛 图片 专业 博客 易生物 视频动画
生命科学实验中心
导航: 生物秀 > 实验 > RNA 实验 > RNA干扰(RNAi) > 正文
  • 倾力提供最优!
  • 生物秀实验频道
Lipid siRNA transfection
作者:佚名 来源:McManus Lab 时间:2007-7-18

    For some cell lines, either electroporation or cationic lipids may be used.  Basically any tfx protocol will require you to work out conditions for every cell line.  Sometimes between 70-90% or more cells can be transfected with cationic lipids.  With a bit of tweaking, and depending on the cell line, you can drive siRNAs into almost 100% of the cells with the following protocol:

     

    Seed cells high, so that they are 85%+ on the day of transfection:

    For one 10 cm dish:

    40 ul Lipofectamine 2000

    20 ug DNA

    100-300 pmoles siRNA

     

    tube 1

     

    mix these two tubes immediately, for a volume of 4 ml, and then add to cells which have been washed 2X with SF-DMEM.  Incubate 3-6 hours, then add normal media.  Check for expression or knockdown 36+ hours post-transfection.  Knockdown by siRNAs usually last 3-5 cell doublings.

     

     

    When oligofectamine is used, the identical protocol is used, but:

    Seed cells at 15% confluency, perform tfx with 30 ul oligofectamine and siRNAs, then repeat tfx 36 hours later.  Analysis is performed when cells are 85-90% confluent.  I have not calculated the transfection efficiency with oligofectamine.

     

     

     

     

     

     

     

     

     

     

关于〖Lipid siRNA transfection〗的最新评论
昵称:      评分: 1分 2分 3分 4分 5分
内容:
生物秀实验频道
设为首页 | 加入收藏 | 关于我们 | 战略伙伴 | 友情链接 | 法律声明 | 广告服务 | 联系我们 | 网站地图
Copyright © 2003-2008 生物秀 (中国·生物秀科技) 版权所有 信产部备案:鲁ICP备05001831号
客服信箱:info@bbioo.com  客服电话:15800302289  客服QQ:254857951
www.bbioo.com All Rights Reserved.