The effects of expression of mutant ASXL1 and HHEX on hematopoietic cells

To identify target genes of mutant ASXL1 (ASXL1-MT) and HHEX in hematopoietic cells, we performed RNA-seq using RUNX1-ETO expressing cord blood cells transduced with vector or ASXL1-MT together with vector or HHEX. Overall design: Method: RUNX1-ETO expressing cord blood cells were transduced with control vector, ASXL1-MT (coexpressing GFP), HHEX (coexpressing NGFR) or ASXL1-MT+HHEX. 48hr after trunsduction, GFP+/NGFR+ cells were sorted by FACS. Total RNA extracted from sorted GFP+/NGFR+ cells were then subjected to RNA-seq analysis. Three biologically independent experiments were perfomed for each sample.

为鉴定造血细胞中突变型ASXL1（ASXL1-MT）与HHEX的靶基因，我们采用转导了对照载体、ASXL1-MT（共表达绿色荧光蛋白GFP）、HHEX（共表达神经生长因子受体NGFR），或ASXL1-MT与HHEX组合载体的、表达RUNX1-ETO的脐带血细胞进行RNA测序（RNA-seq）。实验总体设计：实验方法：将表达RUNX1-ETO的脐带血细胞分别转导至对照载体、ASXL1-MT（共表达GFP）、HHEX（共表达NGFR）或ASXL1-MT与HHEX的组合载体。转导48小时后，通过荧光激活细胞分选（FACS）分选出GFP+/NGFR+双阳性细胞。提取分选得到的双阳性细胞的总RNA，随后进行RNA-seq分析。每个样本均独立重复三次生物学实验。