The important things to remember are
-always use DEPC water (1 mL DEPC / 1 L water, then autoclave for 30 min)
-always bake glassware, utensils, anything not plastic (we bake
overnight at 250F) and autoclave plastic
-always use gloves when handling anything RNase free
-RNase Away is your best friend (but cannot be used on metal)
We use separate reagents for RNase free solutions, but some people
seem to think that doesn't matter.
In regards to starting the RNase free process, when you sacrifice the
embryo, it itself has RNases, so making anything before this RNase
free is futile. We consider our embryos RNase free after they have
been fixed in 4% para (made in DEPC water, in baked bottles, aliquoted
into sterile/new Falcon tubes). After this, every solution used is
Our cryostat is by default RNase free since you have to wear gloves
because it's cold. If you use a microtome that isn't RNase free, just
use a separate blade and wipe down whatever you use with RNase away.
In the case of the glass staining dishes, which can't be baked, we
rinse them with RNase Away and reuse the same dishes for each
experiment. This way they are separated from not RNase free dishes,
and we know they haven't been contaminated in the washing process.
penguins, spoons, and you--what's life like among the flightless?
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