Over the past few decades, the popular scientific view of ribonucleases – aka RNases – has become much richer and more nuanced than it was in the recent past. It has long been understood that RNases are important – important enough for every living thing to have their own suite of them – but it has taken a while for even experts to give RNases credit for being much more than simple destroyers of RNA. The diversity of form and function of these enzymes is only now coming into view, and researchers regularly take new steps in our understanding of how essential RNases are to the everyday activity of all organisms.
When thinking about RNA extraction in the lab, however, RNases still face the reputation they’ve had since the beginning-“RNases are the enemy. And they’re everywhere.”
There’s very little that’s more frustrating than completing an RNA extraction procedure, taking care to follow every step as precisely as possible, only to find that your final yield is many times lower than what was expected. Unfortunately, RNases lurk around every corner in the environment and are more than ready to chop up all that fresh RNA you’re exposing to the world.
So, how should you control for this?
- Make sure your reagents are up to snuff. You’re most likely already doing this but, ensure that all water and buffers are appropriately RNase-free. Reagents that are included with an RNA extraction kit should be fine on this front (unless noted otherwise by the manufacturer), so you mainly need to watch out when bringing “outside” water into the fold. Water treated with diethyl pyrocarbonate (DEPC) is widely available from a variety of sources and is suitable for this purpose. You can also find water that is RNase-free by means other than DEPC treatment. These products may be labeled as “nuclease-free,” indicating that they are also free of DNases (and thus still useful for your RNA extraction purposes).
- Keep your skin away from the reagents. RNases are very common in bodily fluids. Always wear gloves and, if you touch anywhere that is also touched by ungloved hands, change them. In fact, you should feel free to change your gloves often during RNA extraction if you feel like their RNase-free status may have been compromised. The cost of an additional pair of gloves is almost certainly minuscule compared to the cost of the RNA samples you’re working with.
- Mind your surroundings. It isn’t just you. The environment contains a lot of other organisms making RNases of their own, and they’re floating all around you on dust particles as you read this. Try to do your RNA work in a location that doesn’t get a lot of traffic and air flow. Doing work in a hood is ideal.
- Spray the RNases away. There are several products designed to inactivate RNases when applied to surfaces. They can vary in composition and application protocol (be sure to follow manufacturers’ instructions for use), but the goal is the same for all of them: Effectively decontaminate a surface from RNases by eliminating their destructive capabilities. Such products can usually be used on all manner of surfaces, from benchtops to pipettes to gloves themselves, to add an extra layer of security to your RNA extractions.
RNases truly are a category of enzyme whose horizons within our knowledge are constantly expanding. The more we learn about the complex roles of RNA in many aspects of metabolism, the more we recognize the value of trimming that RNA in very specific ways.
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