M*****n
发帖数: 16729 | 1 I need subclone a big promoter, which makes the plasmid over 30 kb.
What kind of bacterial strain should I use to propogate the plasmid and for
subcloning?
What kind of plasmid preparation kit should I use for the plasmid?
Thanks a lot for sharing your experience. |
k*****o
发帖数: 1486 | 2 Only 30kb? Just E. coli. Maybe DH5alpha or others. Anyway, people use it all the time. |
M*****n
发帖数: 16729 | 3 any strain should work?
I will need make a Cre-ER construct, do I need to use a special strain? |
k*****o
发帖数: 1486 | 4 Size doesn't matter, cos a regular cosmid/fosmid would be over 30 kb. Sorry
that I don't know anything about "Cre-ER construct", but I think it's the
protein that you want to express really matters. But not the size. |
C*******e
发帖数: 4348 | 5 Invitrogen 的DH10B是适合做大Plasmid还有cosmid的
不好的是本身有Sp抗性
for
【在 M*****n 的大作中提到】
: I need subclone a big promoter, which makes the plasmid over 30 kb.
: What kind of bacterial strain should I use to propogate the plasmid and for
: subcloning?
: What kind of plasmid preparation kit should I use for the plasmid?
: Thanks a lot for sharing your experience.
|
M*****n
发帖数: 16729 | |
a********k
发帖数: 2273 | 7 不是
【在 M*****n 的大作中提到】
: 这么大的plasmid是不是一定要电转？
|
M*****n
发帖数: 16729 | 8 ok. thanks.
我就先试试heat-shock 转化吧。 |
C*******e
发帖数: 4348 | 9 不是，不过Invitrogen的MegaX DH10B是电转的感受态细胞
挺好用的
【在 M*****n 的大作中提到】
: 这么大的plasmid是不是一定要电转？
|
