A novel phenotypic assay of hepatitis B virus polymerase with extensive site-specific mutagenesis |
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| Authors: | Ya Liu Ying-Ying Luo Xue-Fei Cai Quan-Xin Long Chun-Yang Gan Liu-Qing Yang Haitao Guo Ai-Long Huang Wen-Lu Zhang Jie-Li Hu |
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| Affiliation: | 1.#These authors contributed equally to this work.2.Institute for Viral Hepatitis, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, the Second Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China3.Department of Microbiology and Immunology, Indiana University School of Medicine，Indianapolis 46202, USA4.Collaborative Innovation Center for diagnosis and treatment of infectious diseases (CCID), Hangzhou 310058, China |
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| Abstract: | In summary, we developed a novel mutagenesis phenotypic assay with the following characteristics: (1) acquisition of extensive site-specific mutations through a PCR step and FSR; (2) production of a lentivirus pool expressing different polymerase mutants via one-time lentivirus packaging; (3) generation of stable cell lines that replicate HBV DNA by integrated polymerase mutants, which can help to establish an experimental system with lower variation and higher convenience. This randomized phenotypic assay of HBV polymerase provides a useful tool for assessing the replication fitness and drug susceptibility of HBV variants and quasi-species, which may offer new insight for the NUC therapy and the prognosis of patients with chronic hepatitis B. |
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