Hello,
I performed the total RNA isolation on my samples using miRCURY cell and plant kit, that also helps in preserving miRNAs. The last step of elution, asks me to elute the sample in 50 ul of elution buffer. I then estimated the concentration of my samples, which are too low to be used for further steps. Is there any way of concentrating the RNA eluted in the elution buffer?
Any inputs would be helpful.
Thanks,
You could freeze dry it. When the dry RNA is resuspended in a smaller volume, it will be more concentrated. The other non-volatile substances present in the sample (buffer, salts) will also be concentrated.
Thank you very much Dr. Shapiro. Could you provide me a brief protocol on how to do it? Since, I have never performed this before, I do not want to mess up with my samples. Also, can I resuspend in RNAse free water when scaled down to a small volume?
To lyophilize a sample contained in a 1.5-ml microcentrifuge tube, first freeze the sample. Open the cap. Cover the opening by stretching Parafilm over it. Puncture the Parafilm several times with a small pipette tip. Put the tube in a lyophilizer flask. To keep the top of the tube facing upwards, you should put it in some kind of holder, such as part of a styrofoam 15-ml tube rack. Attach the flask to the lyophilizer and open it to the vacuum. Leave the sample for sufficient time to dry it. Since the volume is only 50 µl, it should only take an hour or two. Let the air back into the flask. Redissolve the dry sample in a smaller volume of RNA-free water.
You could also use a Speed-Vac if there is one available.
Thank you very much. I was told that speed vac would not work if I had my sample resuspended in elution buffer. And performing speed vac could cause significant loss in the yield and integrity of RNA.
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