[Screening and characterization of nucleic acid aptamers specifically binding to adhesin HpaA from Helicobacter...

Objective To screen aptamers that specifically bind to adhesin HpaA from Helicobacter pylori (H. pylori) by systematic evolution of ligands by exponential enrichment (SELEX) and identify the binding properties of aptamers. Methods The prokaryotic expression recombinant plasmid pET28a/HpaA was constructed and the HpaA protein was expressed and purified with IPTG. With the recombinant HpaA protein as target, we screened aptamers with high affinity and specificity binding force by SELEX. The binding force between aptamers and H. pylori in vitro and the performance of aptamers in H. pylori detection from the biopsy of gastric mucosa were examined using the aptamers we had screened. Results We extracted genome from H. pylori ATCC26695 strains and amplified 699 bp HpaA gene using PCR. The recombinant plasmid pET28a/HpaA was constructed successfully. The recombinant HpaA was expressed and purified up to 98% as target for aptamer screening. The six highest affinity aptamers were obtained and named HA1 to HA6 through 10-round positive screening and five-round negative screening by SELEX. The full-length aptamer HA6 and the core sequence of HA6 showed highest affinity and specificity in H. pylori detection in vitro. In view of this, the FAM-labelled aptamer HA6 was used to detect H. pylori in gastric mucosa from 166 patients. The aptamer HA6 showed a higher detection rate (94.58%) than URT (87.95%) in the same batch of clinical samples. Conclusion The aptamers that specifically bind to HpaA may be applied for the detection of H. pylori in gastric mucosa as a novel method.