The HSV-1 ICP22 protein selectively impairs histone repositioning upon Pol II transcription downstream of genes (RNA-seq). The HSV-1 ICP22 protein selectively impairs histone repositioning upon Pol II transcription downstream of genes (RNA-seq)

Primary human fetal foreskin fibroblasts (HFFF) were infected with the ICP22-null mutant of HSV-1 and HSV-1 Wt strain F at a multiplicity of infection (MOI) of 10 for 8 and 12 hpi. Cells were treated for 8 or 12 hours with the phosphonoacetic acid (PAA) (350ug/ml). Total cellular RNA was isolated using Trizol. T-HF cells were grown either in the presence or absence of DOX. Upon DOX exposure, cells expressed either HA-ICP22 or HA-ICP22 and V5-ICP27. T-HF HA-ICP22 cells were treated with 80 mM KCl (salt stress) for 2 hours before collecting the total RNA. Total cellular RNA was isolated using Trizol. Overall design: Primary human fetal foreskin fibroblasts (HFFF) were infected with the ICP22-null mutant of HSV-1 and HSV-1 Wt strain F at a multiplicity of infection (MOI) of 10 for 8 and 12 hpi. Total cellular RNA was isolated using Trizol. T-HF cells were grown either in the presence or absence of DOX. Upon DOX exposure, cells expressed either HA-ICP22 or HA-ICP22 and V5-ICP27. T-HF HA-ICP22 cells were treated with 80 mM KCl (salt stress) for 2 hours before collecting the total RNA. Total cellular RNA was isolated using Trizol.

原代人胎儿包皮成纤维细胞（Primary human fetal foreskin fibroblasts, HFFF）以感染复数（multiplicity of infection, MOI）为10的比例，分别感染单纯疱疹病毒1型（Herpes Simplex Virus 1, HSV-1）ICP22缺失突变株与HSV-1野生型F株，于感染后8小时和12小时（hours post infection, hpi）收取样本。细胞经350 μg/ml的膦甲酸钠（phosphonoacetic acid, PAA）处理8或12小时后，采用Trizol试剂提取总细胞RNA。
T-HF细胞分别在添加或不添加多西环素（Doxycycline, DOX）的培养基中培养。当加入DOX诱导表达后，细胞可表达HA标签融合ICP22（HA-ICP22），或同时表达HA-ICP22与V5标签融合ICP27（V5-ICP27）。将T-HF HA-ICP22细胞以80 mM氯化钾（KCl）进行盐胁迫处理2小时后，收集总细胞RNA，采用Trizol试剂提取总细胞RNA。
实验设计概述：原代人胎儿包皮成纤维细胞（HFFF）以感染复数（MOI）为10的比例，分别感染HSV-1 ICP22缺失突变株与HSV-1野生型F株，于感染后8小时和12小时（hpi）收取样本，采用Trizol试剂提取总细胞RNA。T-HF细胞分别在添加或不添加DOX的培养基中培养。当加入DOX诱导后，细胞可表达HA-ICP22，或同时表达HA-ICP22与V5-ICP27。将T-HF HA-ICP22细胞以80 mM KCl进行盐胁迫处理2小时后，收集总细胞RNA，采用Trizol试剂提取总细胞RNA。