I would use tri-reagent to isolate the RNA. To your cell-free system, spike-in tri-reagent and do a simple RNA extraction. I personally would use the Direct-zol kit from Zymo research, which uses tri-zol in protocol. With that kit you get good quality RNA and can perform in-column DNase treatment.

Kyle Skalenko Thank you for your advice! I am planning to use RNeasy plant kit to extract total RNA. The concerning point is low RNA amount and small-scale volume of my eukaryotic cell-free reactions.

I see. RNeasy is an ok choice.

The yield is a different story. When you purchase a kit, make sure it allows for small volume elution, this way you can concentrate the RNA immediately off the column instead of doing a precipitation step.

What you'll probably have to do is consolidate multiple reactions into one RNA prep. But first step is to determine how much RNA you can get from a single reaction. I also advise to quantify RNA using a qubit.