Expression and methylation array analysis of CD19+ B-cells from Chronic lymhocytic leukemia (CLL) patients and age matched healthy donor samples [Expression]. Homo sapiens

Whole genome sequencing revealed CLL as a disease of the genome and epigenome defined by somatic mutations and aberrant DNA-methylation. To uncover the impact of aberrant methylation on transcription, gene expression and methylation array profiling was performed in CLL and B-cells. RNA from 13 CLL patients and 6 healthy donor samples was analyzed on expression arrays. Overall design: Total RNA was obtained from CD19+ sorted B-cells and CLL cells using the DNA/RNA all prep Kit (Qiagen). Peripheral blood mononuclear cells (PBMCs) were isolated from whole blood by density centrifugation using Biocoll. CD19+ PBMCs were purified using CD19 Micro Beads for magnetic positive selection.

全基因组测序（Whole Genome Sequencing）研究证实，慢性淋巴细胞白血病（Chronic Lymphocytic Leukemia, CLL）是一类由体细胞突变与异常DNA甲基化共同定义的基因组与表观基因组疾病。为探究异常甲基化对转录及基因表达的调控作用，本研究针对慢性淋巴细胞白血病细胞与B细胞开展了甲基化芯片（Methylation Array）分析；同时针对13例慢性淋巴细胞白血病患者与6例健康供体的样本提取RNA，通过表达芯片（Expression Array）完成基因表达水平检测。
实验整体设计：
总RNA提取采用DNA/RNA总纯化试剂盒（DNA/RNA all prep Kit，Qiagen），从经分选的CD19阳性（CD19+）B细胞与慢性淋巴细胞白血病细胞中获取总RNA。采用Biocoll分层离心液通过密度梯度离心法从全血中分离外周血单个核细胞（Peripheral Blood Mononuclear Cells, PBMCs）。通过CD19微珠（CD19 Micro Beads）进行磁珠阳性分选，以纯化CD19阳性外周血单个核细胞。