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Cloning of ACE-2 gene encoding the functional receptor for the SARS coronavirus and its expression in eukaryotic cells

Lu, J.; Zhou, B-Ping.; Wen, L-Xia.; Jiang, X-Ling.

Zhonghua Shi Yan He Lin Chuang Bing du Xue Za Zhi 19(3): 260-263

2005

ISSN/ISBN: 1003-9279
PMID: 16261211
Document Number: 590933
Total RNA was extracted from the right atrial tissue of a patient with heart failure, and the full-length angiotensin converting enzyme 2 (ACE-2) gene was amplified by a nested reverse transcription-polymerase chain reaction method. The ACE-2 gene was cloned into the pcDNA4/HisMax-TOPO eukaryotic expression vector to construct the recombinant plasmid pcDNA4/ACE-2, and then transfected into 293 T lymphocytes. ACE-2 transient expression was examined by western blotting. Syncytia inhibition assay was developed to detect neutralizing antibodies against severe acute respiratory syndrome coronavirus (SARS-CoV), and was compared with SARS pseudovirus neutralization assay. The recombinant plasmid pcDNA4/ACE-2 expressed ACE-2 in eukaryotic cells and induced cell fusion between the S protein- and ACE-2-expressing cells. This cell-cell fusion assay was used to detect SARS-CoV neutralizing antibodies.

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