Abstract: With a RT-PCR method we cloned a drought tolerant DHN2b gene (designated PdDHN2b) from Populus deltoides×Populus nigra. The open reading frame of PdDHN2b is 1 440 bp and encodes 379 amino acids of which most are hydrophilic, without an obvious transmembrane domain. A subcellular localization assay showed that PdDHN2b is localized in the membrane orcytoplasm. Real time fluorescence quota PCR expression analysis suggests that PdDHN2b is strongly expressed in top leaves, but not in functional leaves, roots or stems. PdDHN2b is induced by NaCl, ABA and drought. A recombinant pET-PdDHN2b vector was constructed for prokaryotic expression and expressed in E. coli BL21. Fusion proteins were purified and used to immunize white mice to obtain antiserum. SDS-PAGE electrophoresis showed that the molecular weight of the fusion protein was 58 ku. In the end, we used the Ni-NTA purified recombinant PdDHN2b-his protein for Western blot analysis.