v***a
发帖数: 1242 | 1 transfection的efficiency是否跟cell density相关呢？细胞少了是不是efficiency就
高，但是toxicity会大？
我做plasmid DNA transfection的时候，发现普通control（没有加任何transfection
reagent的）和有vector plasmid transfection的control相比，我要观测的蛋白表达
不一样，有vector的蛋白少了很多。不知道这是怎么回事呢？
谢谢～ |
n********k
发帖数: 2818 | 2 it is not a proper comparison, many transfection reagents impact on cells...
think about it, they are peneratring
cell membranes...
transfection
【在 v***a 的大作中提到】
: transfection的efficiency是否跟cell density相关呢？细胞少了是不是efficiency就
: 高，但是toxicity会大？
: 我做plasmid DNA transfection的时候，发现普通control（没有加任何transfection
: reagent的）和有vector plasmid transfection的control相比，我要观测的蛋白表达
: 不一样，有vector的蛋白少了很多。不知道这是怎么回事呢？
: 谢谢～
|
v***a
发帖数: 1242 | 3 嗯，加了transfection reagent的control某些指标明显跟未加reagent的control不一
样了。那我在paper里可以这样说明吗？我老板说应该保持前后一致。比如我前面一个
蛋白在无transfection的cells里表达量有较多，但现在在有vector的control里，蛋白
的表达量下降了好多，用RT-PCR检测几乎就要看不到了；老板就要我重做，说这2个
control非得match。大家遇到过这样的情况吗？如果前后不match，该如何说明比较好？
..
【在 n********k 的大作中提到】
: it is not a proper comparison, many transfection reagents impact on cells...
: think about it, they are peneratring
: cell membranes...
:
: transfection
|
c**n
发帖数: 73 | 4 You are supposed to transfect cells with empty vector as a control
好？
【在 v***a 的大作中提到】
: 嗯，加了transfection reagent的control某些指标明显跟未加reagent的control不一
: 样了。那我在paper里可以这样说明吗？我老板说应该保持前后一致。比如我前面一个
: 蛋白在无transfection的cells里表达量有较多，但现在在有vector的control里，蛋白
: 的表达量下降了好多，用RT-PCR检测几乎就要看不到了；老板就要我重做，说这2个
: control非得match。大家遇到过这样的情况吗？如果前后不match，该如何说明比较好？
:
: ..
|
v***a
发帖数: 1242 | 5 Yes, I did. But my boss required that the result from empty vector control
be the same as the normal control to keep the consistence... How can I
explain it solidly enough to convince my boss? Or in fact I did something
wrong that the influence of the empty vector control should not be that much
(made the mRNA level of my protein nearly undetectable)?
【在 c**n 的大作中提到】
: You are supposed to transfect cells with empty vector as a control
:
: 好？
|
n********k
发帖数: 2818 | 6 your boss is typically those ...I don;t know what to say...BTW, the level of
mRNA etc we normally detect is
always in a relative sense, I don't see either you or your boss has a point
here..., so just do another control to
see whether it is your vector or transfection reagent problems: add
transfections reagents without any DNA or
different vectors...BTW, No offense, I suppose you are a newbie, otherwise u
got quite some work to do with
your design and interpretation of experiments and troublin
【在 v***a 的大作中提到】
: Yes, I did. But my boss required that the result from empty vector control
: be the same as the normal control to keep the consistence... How can I
: explain it solidly enough to convince my boss? Or in fact I did something
: wrong that the influence of the empty vector control should not be that much
: (made the mRNA level of my protein nearly undetectable)?
|
g*****y
发帖数: 6325 | 7 cell density在70%-80%的transfection的efficiency 最高
transfection
【在 v***a 的大作中提到】
: transfection的efficiency是否跟cell density相关呢？细胞少了是不是efficiency就
: 高，但是toxicity会大？
: 我做plasmid DNA transfection的时候，发现普通control（没有加任何transfection
: reagent的）和有vector plasmid transfection的control相比，我要观测的蛋白表达
: 不一样，有vector的蛋白少了很多。不知道这是怎么回事呢？
: 谢谢～
|
v***a
发帖数: 1242 | 8 谢谢！！！我承认你说得都对，I've got a long, long way to go
of
point
u
【在 n********k 的大作中提到】
: your boss is typically those ...I don;t know what to say...BTW, the level of
: mRNA etc we normally detect is
: always in a relative sense, I don't see either you or your boss has a point
: here..., so just do another control to
: see whether it is your vector or transfection reagent problems: add
: transfections reagents without any DNA or
: different vectors...BTW, No offense, I suppose you are a newbie, otherwise u
: got quite some work to do with
: your design and interpretation of experiments and troublin
|
|
