Transcription profiling of murine presomitic mesoderms of 17 samples at various time points to identify cyclic genes of the mouse segmentation clock

A microarray time series was generated to identify cyclic genes of the segmentation clock in the mouse. The right posterior half presomitic mesoderms (PSM) from 17 mouse embryos were dissected while the contralateral side of the embryo containing the left PSM was immediately fixed to be analyzed by in situ hybridization using a Lfng probe to order the samples along the segmentation clock oscillation cycle. Probes were produced from RNA extracted from the 17 dissected posterior half PSMs using a two-step amplification protocol and were hybridized to Affymetrix GeneChip MOE430A. The reproducibility of the amplification procedure was initially assessed by comparing array data generated from the right and the left posterior PSM from the same embryo. Because of the symmetry of the paraxial mesoderm along the left-right axis, left and right samples are expected to show overtly similar gene expression. RNA was amplified from three such sample pairs (1, a and b; 2, a and b; 3, a and b) and hybridized on Murine Genome U74Av2 array (MG-U74Av2)本研究生成微阵列时间序列数据集，用于鉴定小鼠分节时钟（segmentation clock）的周期基因。我们从17只小鼠胚胎中分离出右侧后半体节前中胚层（presomitic mesoderm, PSM），同时将携带左侧PSM的胚胎对侧组织立即固定，随后使用Lfng探针开展原位杂交（in situ hybridization）分析，以依据分节时钟的振荡周期对样本进行排序。探针通过两步扩增流程，从17份分离得到的后半PSM样本提取的RNA中制备，并与Affymetrix GeneChip MOE430A芯片完成杂交。本研究首先通过比对同一胚胎的右侧与左侧后半PSM的芯片数据，评估了扩增流程的可重复性。鉴于轴旁中胚层沿左右轴具备对称性，理论上左右两侧样本的基因表达谱应呈现高度相似性。我们从3组此类样本对（1组a与b；2组a与b；3组a与b）中扩增得到RNA，并将其与小鼠基因组U74Av2芯片（MG-U74Av2）进行杂交。