MACHEREY-NAGEL – 05 / 2014, Rev. 05
12
5 Protocols
5.1 RNA Clean-up
Before starting the preparation:
• Check if Wash Buffer RA3 was prepared according to section 3.
1 Sample preparation
Fill up RNA samples smaller than 100 μL with RNase-free
water to
100 μL
.
RNA samples from 100–200 μL should be filled up with
RNase-free water to 200 μL.
Fill up RNA
sample to
100 μL with
water
2 Preparation of lysis-binding buffer premix
Prepare a Buffer RA1-ethanol premix with a ratio of 1:1.
For each
100 μL RNA sample
mix
300 μL Buffer RA1
and
300 μL of ethanol (96–100 %)
.
If multiple samples are processed, the preparation of a
master-premix is recommended (e.g., 2 mL Buffer RA1 +
2 mL 98 % ethanol for approximately 6 preparations).
Prepare
premix:
Mix
300 μL RA1
with
300 μL ethanol
(96–100 %)
3 Adjust RNA binding conditions
To
100 μL RNA sample
add
600 μL (6 volumes) of
Buffer RA1-ethanol-premix. Mix sample with premix by
vortexing.
If a 200 μL RNA sample is processed, add 1200 μL
Buffer RA1-ethanol premix.
After addition of ethanol a stringy precipitate may become
visible which will not affect the RNA isolation. Be sure to mix
thouroughly and apply sample as homogeneous solution
onto the column.
+ 6 vol.
premix
Mix
NucleoSpin
®
RNA Clean-up
