v****e
发帖数: 131 | 1 有人做过siRNA和plasmid cotransfection吗？我用2000转的不好。可是分别单转都还
好。有什么经验可以分享的吗？thanks! |
h******y
发帖数: 351 | 2 a couple of suggestions.
1. optimize the ratio of your siRNA and plasmid DNA, as they will compete fo
r lipo-2000
2. try other non-liposome based reagents, such as FuGene 6 or PEI.
【在 v****e 的大作中提到】
: 有人做过siRNA和plasmid cotransfection吗？我用2000转的不好。可是分别单转都还
: 好。有什么经验可以分享的吗？thanks!
|
p*****e
发帖数: 280 | 3 Thanks for your suggestions, LZ's trouble is also mine.
BTW, what's the ideal ratio for siRNA/plasmid?
fo
【在 h******y 的大作中提到】
: a couple of suggestions.
: 1. optimize the ratio of your siRNA and plasmid DNA, as they will compete fo
: r lipo-2000
: 2. try other non-liposome based reagents, such as FuGene 6 or PEI.
|
b****s
发帖数: 148 | 4 40pmol RNAi vs. 1ug dsDNA worked fine for me.
i used lipo2000
【在 p*****e 的大作中提到】
: Thanks for your suggestions, LZ's trouble is also mine.
: BTW, what's the ideal ratio for siRNA/plasmid?
:
: fo
|
w********r
发帖数: 1431 | 5 分别转，lipo2000转质粒的效率在我的手里效率比较差（也可能我们的细胞比较不容转
染）。
siRNA和lipoRNAiMax在一个管子里混好，
DNA和lipofectimine或者lipoLTX在另一个管子里混好，
然后一起加进去。
因为两种东西一起加毒性会大一点，细胞会死一些，
所以转染的时候，细胞的起始密度可以比单独转染siRNA或者质粒的密度要高一点 |
h******y
发帖数: 351 | 6 calculate the moles of your plasmid DNA based on its size. Then try several
different ratios of your siRNA and DNA, for example, 1:1, 1:2, 2:1, etc.
【在 p*****e 的大作中提到】
: Thanks for your suggestions, LZ's trouble is also mine.
: BTW, what's the ideal ratio for siRNA/plasmid?
:
: fo
|
g*********5
发帖数: 2533 | 7 2000 is toxic for plasmid when cotransfect!
us reagent from mirus. |
m***0
发帖数: 7 | 8 试过用electroporation,效果非常不错 |
