the try cell plus RNA purification kit
is based on try cell reagent lysis
technology and pure links silica
membrane purification technology this
combination provides highly pure RNA
even from difficult samples like fibrous
tissue for your protection
always wear a lab coat gloves and safety
glasses prepare your work area
and wash buffer and gather your
materials prior to starting the
procedure to get started prepare your
lace8 you can start with 50 to 100
micrograms of fresh or frozen tissue or
cells
add one mil of trysil reagent
homogenized thoroughly using a tissue or
rotor stator homogenizer let the sample
stand at room temperature for five
minutes next add point two mils of
chloroform and shake the tube vigorously
for about 15 seconds
then let the tube sit for two to three
minutes at room temperature
centrifuge your sample at 12,000 G for
15 minutes at 4 degrees Celsius
you will see three different layers in
the tube the colorless upper layer
contains the RNA transfer point for
microliters of the upper layer - a fresh
rnase free tube and add an equal volume
of 70% ethanol mix well by vortexing
next run your isolated RNA solution
through a pure link silica membrane
column provided in the kit to wash away
contaminants transfer 700 microliters of
the sample to a column which has been
inserted into a - milk collection tube
centrifuge at 12,000 g for 15 seconds
discard the flow-through wash your bound
RNA by adding seven hundred microliters
of wash buffer one and centrifuging at
12,000 g for 15 seconds discard the
flow-through and the collection tube
with the silica membrane column in a
fresh collection to add 500 microliters
of prepared wash buffer to and
centrifuge at 12,000 g for 15 seconds
discard the flow-through and repeat dry
the silica membrane by centrifuging the
column at 12,000 g for one minute
discard the collection tube to elute
your RNA from the silica membrane insert
the column into the recovery tube then
add the appropriate volume of elution
buffer based on your starting amount
incubate at room temperature for one
minute next centrifuge the column insert
it into a recovery tube at 12,000 g for
two minutes your ultra pure RNA is in
the recovery tube ready to be used or
stored at minus 20 degrees Celsius
you
