Abstract
Nucleos(t)ide analogues (NUCs) susceptibility assay is important for the study of hepatitis B virus (HBV) drug resistance. The purpose of susceptibility assay is to test the sensitivity of a specific HBV variant to NUCs in vitro, by which assesses if and to what extent the mutant virus is resistant to a specific NUC. Among the existing susceptibility assay methods, stable cell line expressing the specific variant is one of the commonly used assessment systems based on its high repeatability. However, establishment of stable cell lines expressing individual variant is laborious and time-consuming. In the present study, we developed a novel strategy for rapidly establishing HBV replicating stable cell lines. We first established an acceptor cell line stably transfected with a polymerase-null HBV 1.1mer genome DNA, then lentiviruses expressing different mutant HBV polymerases were transduced into the acceptor cell line respectively. Stable cell lines replicating HBV DNA with the trans-complemented HBV polymerases were established by antibiotics selection. Lamivudine and entecavir susceptibility data from these polymerase-complementing cell lines were validated by comparing with other assays. Taken together, this transcomplementation strategy for establishment of stable cell lines replicating HBV DNA with clinically isolated HBV polymerase provides a new tool for NUC susceptibility assay of HBV.
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