w******e
发帖数: 1187 | 1 which enzyme should I use to add NTP to 3' end?
can you recommend a protocol too?
thank you! |
a***e
发帖数: 1010 | 2 if the 5'-end of your ssRNA is -OH, you can add a Pi by T4 PNK.
although your substrate is gamma-Pi from ATP, the product is alpha-Pi. |
w******e
发帖数: 1187 | 3 thx, but my 5'end would be occupied by a biotin-GMP already, so
I need to use 3'end:(
【在 a***e 的大作中提到】
: if the 5'-end of your ssRNA is -OH, you can add a Pi by T4 PNK.
: although your substrate is gamma-Pi from ATP, the product is alpha-Pi.
|
a****k
发帖数: 1130 | 4 T4 RNA ligase with 32pCp?
【在 w******e 的大作中提到】
: thx, but my 5'end would be occupied by a biotin-GMP already, so
: I need to use 3'end:(
|
a***e
发帖数: 1010 | 5 yeah, correct.
I struggle some time. it just doesn't jump out of my head.
【在 a****k 的大作中提到】
: T4 RNA ligase with 32pCp?
|
a****k
发帖数: 1130 | 6 the labeling is pretty good in our hand
here's the protocol from ambion
http://www.ambion.com/techlib/misc/RNA3_labeling.html
【在 a***e 的大作中提到】
: yeah, correct.
: I struggle some time. it just doesn't jump out of my head.
|
w******e
发帖数: 1187 | 7 thanks a lot!
【在 a****k 的大作中提到】
: the labeling is pretty good in our hand
: here's the protocol from ambion
: http://www.ambion.com/techlib/misc/RNA3_labeling.html
|
w******e
发帖数: 1187 | 8 btw, did any of you guys try adding alpha ATP using E coli poly A
polymerase? I saw it in a paper once and already bought the stuff,
don't want to waste them...
thanks a lot!
【在 w******e 的大作中提到】
: thanks a lot!
|
