Fig. 6: Aphidicolin and cyclin-dependent kinase inhibitor p21 peptide specifically inhibit the repair of the rcDNA plus-strand.

a Schematic of time course experiments to test the effects of aphidicolin and p21 peptides on repair of HBV RrcDNA plus- and minus-strand repair. b A simplified schematic depicting the four fragments in unrepaired RrcDNA digested by AatII/PsiI. c cccDNA formation of the RrcDNA substrate under treatments of mock (1% DMSO, lanes 1–7) or aphidicolin (100 μM in 1% DMSO, lanes 8–14) was detected by EtBr-containing agarose gel. d–h Repair of plus-strand fragments Pa, Pb, and minus-strand fragments Ma, Mb was monitored by Southern blot d–g or streptavidin blot h. i Repair efficiency of plus-strand e and minus-strand f was calculated and plotted as in Fig. 2i. (−) and (+) repair denote repair of minus and plus-strands, respectively. M, marker. j–p, same as c–i, except that AAA mutant p21 peptide was used as mock treatment, and WT p21 peptide was used as inhibitor to inhibit plus-strand synthesis. All experiments were repeated thrice, and each individual measurement is plotted. The lines connect the average values of three measurements at each time point. Statistical analyses between the repair efficiencies of the minus strand at each time point are performed by two-stage step-up t test method of Benjamini, Krieger, and Yekutieli from Graphpad Prism. P values are 0.0004, 0.000001, 0.0000004, 0.00003, 0.00006, and 0.007 for indicated time points in i. P values are 0.00007, 0.00007, 0.00007, 0.0001, 0.00005, and 0.00005 for indicated time points in p, and statistically significant p values are indicated by “*”. Rrc, RrcDNA; rL, recombinant linear RrcDNA, ccc, cccDNA. Source data are provided as a Source Data file.