Selection of nucleotide probes, future aptamers
Stéphane Octave, a research engineer at UTC’s GEC laboratory, is working with Séverine Padiolleau on Lyme disease. He explains the process of selecting oligonucleotide probes.
Concretely? “First, we use single-stranded DNA or RNA that can be structured and thus acquire the ability to specifically recognise a molecular target. In this case, this oligonucleotide sequence becomes an aptamer,” he says.
The target selection process? “It involves producing the protein of interest in a bacterium other than the one involved in Lyme disease, purifying it and ensuring that it is in its natural conformation so that selection can begin. We attach this protein to magnetic beads, which we then bring into contact with our DNA bank containing 1014 different oligonucleotides. Next, we need to isolate, from among these 1014 DNA strands, those capable of recognising the target and which could become aptamer candidates. This is where the expertise of the laboratory, and in particular that of Prof. Bérangère Bihan-Avalle, comes into play, enabling this selection process to be carried out successfully. If I dared to use a metaphor, I would say that this process is based on the principle of musical chairs, but on a molecular scale. At each selection round, we will collect the candidates that interest us. So, if we attach a protein to a bead and bring our DNA bank into contact with it, a certain number of these DNA molecules will interact, bind to the protein and be collected to make copies for the next round. The rest of the DNA, which is not bound, will be of no interest to us. In the next round, the selection conditions are modified to make it increasingly difficult to bind to our target. In this case, we refer to selection pressure. In this way, we gradually reduce the diversity and obtain a few molecules as candidates qualified for aptamer status. Finally, we synthesise them independently and test them one by one to determine their ability to specifically recognise our target. Of course, we keep the best ones based on a performance threshold compatible with the end-use objective,” explains Stéphane Octave.
MSD
