*2.2. Plasmids and Transfection*

A triple FLAG ® (DYKDDDDK)-tagged ZIKV capsid expression construct was generated by polymerase chain reaction (PCR) using a mammalian expression vector encoding the capsid protein [28] as template. The resulting PCR product was then cloned into pcDNA 3.1(-) plasmid using the restriction sites NheI and XhoI. pCMV3-PEX11B was purchased from Sino Biological Inc. Oligonucleotide primers were designed to amplify the desired PEX11B sequence and introduce a myc epitope tag cassette into the 5 end of the cDNA. The resulting PCR product was then cloned into the lentiviral vector pTRIP-MCS-IRES-AcGFP [30]. All oligonucleotide primers used in this study are shown in Table S1.

For indirect immunofluorescence analysis in U251 cells, transfection of the appropriate expression plasmids was performed using Lipofectamine 2000 (Invitrogen; Carlsbad, CA). Poly(I:C) (Sigma-Aldrich; St. Louis, MO, USA) was transfected into cells using TransIT-LT1 (Mirus Bio; Madison, WI, USA).

#### *2.3. Production of Lentiviral Particles and Transduction of Cells*

Pseudotyped lentiviruses were recovered from the media of HEK293T cells transfected with pTRIP-AcGFP plasmids encoding flavivirus capsids or PEX11B, and titered as described [30]. Transduction of U251 cells with recombinant lentiviruses was performed in DMEM containing 3% FBS and 5 μg/mL polybrene for 48 h in 12-well plates.
