FIG 2.

HPgV protein detection in human astrocytes following infection. (A) The HPgV ΔNS2 mutant was characterized by an 87-nucleotide deletion located in the NS2 gene. The presence of the NS2 deletion was observed by gel electrophoresis following qPCR compared to cDNA prepared from HPgV WT RNA. (B) To verify the presence of a deletion, viral cDNAs created from WT and ΔNS2 plasmids were analyzed by Sanger sequencing. (C) Using HPgV WT (GenBank accession no. AF121950) and ΔNS2 sequences from LE-1 (GenBank accession no. MH179063), a phylogenetic tree was constructed for the mutation site in NS2. (D) Astrocytes were infected with HPgV WT or ΔNS2 (MOI, 0.1) for 7 days before lysates were harvested and immunoblotted for HPgV NS5A and E2 as indicated. This experiment was repeated in two separate biological donors.