B8
发帖数: 92 | 1 最近准备做rnaseq看基因表达的差别，
小鼠的组织样品。
请问各位哪个kit比较好用？
Qiagen的Rneasy kit如何？
提完rna是否还需要dnase处理？还是直接就可以做library了？
谢谢！ |
l********e
发帖数: 415 | 2 Rneasy Kits worked fine for us.
You do need to remove DNA as they can be incorporated into the sequencing
library.
However, DO NOT perform the standard on-column DNase treatment as it lead to
dramatic sample loss and often times not complete. I would just Dnase the
sample after purification and heat inactivate.
Please note that the regular Rneasy columns do not work well with RNA <
200bp. Therefore, if you want to look at small RNA, you might need a special
kit. |
D*a
发帖数: 6830 | |
B8
发帖数: 92 | 4 facility说的很不清楚。
【在 D*a 的大作中提到】
: 找哪个facility做就问他们不就行了
|
n***w
发帖数: 2405 | 5 We work with tiny mouse tissues and we use RNeasy plus micro kit.
Just followed the instructions and worked fine for RNA-seq.
Qiagen has so many kits designed for so many tissue types. Check their
website. |
v********a
发帖数: 646 | 6
The RNeasy Plus Mini Kit purifies up to 100 μg total RNA (>200 nt) from a
single extraction with efficient gDNA removal.
RNAs over 200nt are eligible for RNA-seq?
【在 n***w 的大作中提到】
: We work with tiny mouse tissues and we use RNeasy plus micro kit.
: Just followed the instructions and worked fine for RNA-seq.
: Qiagen has so many kits designed for so many tissue types. Check their
: website.
|
B8
发帖数: 92 | 7 提完RNA后，做dnase treatment吗？
kit说一般不需要，
网上一些信息和前面的一个大侠都做的。
谢谢。
【在 n***w 的大作中提到】
: We work with tiny mouse tissues and we use RNeasy plus micro kit.
: Just followed the instructions and worked fine for RNA-seq.
: Qiagen has so many kits designed for so many tissue types. Check their
: website.
|
D*a
发帖数: 6830 | 8 那就问问都给谁做过，过去问问。
【在 B8 的大作中提到】
: facility说的很不清楚。
|
B8
发帖数: 92 | 9 这个KIT主要是不能bind小的RNA
【在 v********a 的大作中提到】
:
: The RNeasy Plus Mini Kit purifies up to 100 μg total RNA (>200 nt) from a
: single extraction with efficient gDNA removal.
: RNAs over 200nt are eligible for RNA-seq?
|
i*********0
发帖数: 915 | 10 如果不是微量的组织的话，可以考虑用，Tri-reagent.
RNA 抽提完了以后，跑个 胶看看有没有明显的降解，如果没有的话，可以直接交给
facility了。 |
B8
发帖数: 92 | 11 谢谢，我们自己做library的，
给facility都是马上就可以测序的。
【在 i*********0 的大作中提到】
: 如果不是微量的组织的话，可以考虑用，Tri-reagent.
: RNA 抽提完了以后，跑个 胶看看有没有明显的降解，如果没有的话，可以直接交给
: facility了。
|
a******n
发帖数: 45 | 12 如果RNA量大的话， trizol 》DNase I treatment 》Qiagen RNeasy Mini Kit
purify。 量小的话，RNeasy plus micro kit或者Life Arcturus Picopure RNA
isolation Kit， on column DNase digestion。不要直接用trizol提的RNA做library
，纯度不够。 |
B8
发帖数: 92 | 13 我们量小，那就准备用rneasy micro kit.
前面一位大侠不建议做on column digestion. 想知道有没办法看一下是否把DNA除了？
我们可以跑bioanalyzer.
谢谢
library
【在 a******n 的大作中提到】
: 如果RNA量大的话， trizol 》DNase I treatment 》Qiagen RNeasy Mini Kit
: purify。 量小的话，RNeasy plus micro kit或者Life Arcturus Picopure RNA
: isolation Kit， on column DNase digestion。不要直接用trizol提的RNA做library
: ，纯度不够。
|
