<p>a) Sensitivity of DNA cassette to DNase I. DNA cassette complexed with lipofectamine and DNA nanocapsules were incubated for 1 hour without DNase I and with DNase I, respectively. b) Viability of HEK-293T cells transduced with CCR5 DNA cassette nanocapsules. HEK-293T cells were treated with DNA cassette nanocapsules at 0, 0.1, 0.2 and 0.4 pmol for 4 h at 37°C in 100uL of serum-free medium. After 24 h, cell viability was determined with CytoToxGlo kit using a 96-well plate reader. c) Knockdown of CCR5-Luciferase in HEK-293T cells by CCR5 DNA cassette nanocapsules and CCR5 siRNA lipofectamine complex. d) Down-regulation of CCR5 in 293 cells by sh1005 DNA cassette nanocapsules with different ratios (5:0; 3:2; 2:3 and 1:4) of degradable crosslinker (Glycerol 1,3-diglycerolate diacrylate, GDGDA) to non-degradable crosslinker (N,N’-methylenesbisacrylamide, BIS). On day 0, cells were transduced with DNA cassette nanocapsules for 4 hours. On day 3, 5 and 9, cells were stained and analyzed by flow cytometry. e) Inhibition of viral infection of Affinofile cells by sh1005 DNA cassette nanocapsules with different ratios (5:0; 3:2; 2:3, 1:4 and mixture of 5:0 (50%) and 2:3 (50%)) of degradable crosslinker (GDGDA) to non-degradable crosslinker (BIS). On day 0, Affinofile cells were transduced with DNA cassette nanocapsules for 4 hours and then cultured in the induction medium (6 ng/ml doxy and 5 mM ponA, respectively). On day 3, 5 and 9, Affinofile cells were seeded into 96-well plates at a density of 10^4 cells/well. 24 hours later, the induction medium was removed and gently replaced with 100ul of fresh, warmed culture medium containing env-pseudotyped virus. The infection plates were spinoculated at 2,000 rpm for 2 hours at 37°C, and then incubated for an additional 48 hours at 37°C. Infection medium was then removed, the cells were lysed, and luciferase activity was assayed using the plate reader.</p
