b******r
发帖数: 111 | 1 I am doing a knockin project. I have done constructs which contain Neo and
HSV-TK for selection in ES cells.
How do I know Neo and HSV-TK in my plasmid work? Specificly,
Do Neo and HSV-TK genes in my plasmids express their proteins in E.Coli? I
tested in this way: grow 4 tubes
of E.Coli, when adding G418, E.Coli can't grow; When adding gancyclovir, E.
Cori grow well. That means Neo
and HSV-TK genes of my plasmids don't express in E.Coli, right? Then why
do Neo and HSV-TK express in
ES cells? I am not sure now. I am treating ES cells with G418 and
gancyclovir.
The sequence of Neo and HSV-TK is right.
thanks | b******r
发帖数: 111 | 2 Anyone knows the answer?
I
【在 b******r 的大作中提到】
: I am doing a knockin project. I have done constructs which contain Neo and
: HSV-TK for selection in ES cells.
: How do I know Neo and HSV-TK in my plasmid work? Specificly,
: Do Neo and HSV-TK genes in my plasmids express their proteins in E.Coli? I
: tested in this way: grow 4 tubes
: of E.Coli, when adding G418, E.Coli can't grow; When adding gancyclovir, E.
: Cori grow well. That means Neo
: and HSV-TK genes of my plasmids don't express in E.Coli, right? Then why
: do Neo and HSV-TK express in
: ES cells? I am not sure now. I am treating ES cells with G418 and
| n********k
发帖数: 2818 | 3 ES is eu while bacteria is pro...they use different promoter systems...If it
is a confirmed vector, you don't have to worry about the issues...or you
just need to sequence to confirm...if I am not mistaking, you are too wary.
..,
I
【在 b******r 的大作中提到】
: I am doing a knockin project. I have done constructs which contain Neo and
: HSV-TK for selection in ES cells.
: How do I know Neo and HSV-TK in my plasmid work? Specificly,
: Do Neo and HSV-TK genes in my plasmids express their proteins in E.Coli? I
: tested in this way: grow 4 tubes
: of E.Coli, when adding G418, E.Coli can't grow; When adding gancyclovir, E.
: Cori grow well. That means Neo
: and HSV-TK genes of my plasmids don't express in E.Coli, right? Then why
: do Neo and HSV-TK express in
: ES cells? I am not sure now. I am treating ES cells with G418 and
| s********r
发帖数: 312 | 4 promoter不一样。测序可以，把几个接头的地方测一下OK就直接转ES了 | b******r
发帖数: 111 | 5 Thank you both
【在 s********r 的大作中提到】
: promoter不一样。测序可以，把几个接头的地方测一下OK就直接转ES了
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