Nuclear and cellular expression data from the whole 16-cell stage Arabidopsis thaliana embryo

Due to its inaccessibility, transgenic methods have been recently developed to isolate tissue- or cell-specific nuclear RNA from the Arabidopsis embryo for transcriptomic profiling. While these approaches have made the it possible to conduct transcriptomic studies at the cellular level, only nuclear and not whole celluar RNA can be isolated. The question thus arrises if nuclear RNA is a reasonable proxy for whole cellular mRNA. To answer this, microarray-based transcriptomic profiling was used to determine genome-wide expression in nuclei and cells isolated from whole 16-cell stage Arabidopsis embryos. We manually pollinated flowers and either isolated whole embryos (manually using an micromanipulator; cEMB) or the nuclei from embryos (using the INTACT method and a whole embryo marker; nEMB) at 16-cell stage (16C) of development (72h after pollination). Following RNA isolation, microarray-based transcriptomic profiling was performed on 3 biological replicates each.

鉴于拟南芥胚胎组分分离难度较高，近年来研究者开发出多种转基因方法，用于从拟南芥胚胎中分离组织或细胞特异性核RNA，以开展转录组分析。尽管此类方法已实现细胞水平的转录组研究，但仅能分离得到核RNA，而非全细胞RNA。由此引发核心疑问：核RNA是否可作为全细胞mRNA的合理替代指标？为解答该问题，本研究采用基于微阵列（microarray）的转录组分析技术，对从完整16细胞期拟南芥胚胎中分离得到的细胞核与细胞进行全基因组表达水平检测。我们对拟南芥花进行人工授粉，于发育至16细胞期（授粉后72小时，简称16C）时，分别通过两种方式获取实验样本：一是利用显微操作仪手动分离完整胚胎（命名为cEMB），二是采用INTACT方法结合全胚胎标记物分离胚胎细胞核（命名为nEMB）。完成RNA提取后，对每组样本设置3次生物学重复，并开展基于微阵列的转录组分析。