Kinetic Screening of Nuclease Activity using Nucleic Acid Probes

This protocol provides a powerful alternative for screening nuclease activity as biomarker of disease, with an easy-to-implement methodology even for researchers who are not as specialized in nucleic acid probes. The main advantage of this technique is the ability to select nucleic acid probes that can identify both known and unknown nuclease activities, taking advantage of the probe-nuclease dynamic interaction. Other advantages of this methodology are its flexibility, high reproducibility and ease of use.

The demonstration will be performed by Khadija, a Masters student, and Baris, a postdoc from our laboratory. When designing an oligonucleotide library, include at least one DNA and one RNA random sequence containing a combination of adenine, guanine, cytosine and thiamine or uracil. To prepare the oligonucleotide probes, spin down the lyophilized oligonucleotide probes and dilute each probe in Tris-EDTA buffer at a 500 picomolar per microliter concentration to prevent nuclease degradation.