Polysomes from DENR knockdown cells. Drosophila melanogaster

The aim was to identify transcripts that are poorly translated upon knockdown of DENR. Lysates from control (GFP) and DENR knockdown S2 cells were run on polysome gradients. RNA from the (80S peak + polysome peaks) was isolated and analyzed by microarrays. In parallel, total RNA from the cells was also profiled as a normalization control and profiled. The 'translation score' of translated mRNA to total RNA was then calculated for control and DENR knockdown cells. Overall design: Two biological replicates for DENR knockdown and GFP knockdown were performed. For each sample, (80S+polysomes) and total RNA were profiled, leading to 8 samples in total.

本研究旨在鉴定DENR基因敲低后翻译效率低下的转录本。以对照（GFP）组与DENR基因敲低的S2细胞制备裂解液，通过多聚核糖体梯度离心进行分离。提取(80S峰与多聚核糖体峰)中的RNA，通过微阵列（microarrays）进行分析检测。与此同时，同步提取细胞总RNA并开展转录组谱分析，以作为标准化对照。随后分别计算对照组与DENR基因敲低组细胞中，翻译态mRNA与总RNA的“翻译评分”。实验整体设计：设置DENR基因敲低组与GFP基因敲低组，每组均完成2次生物学重复。每个样本均分别对(80S+多聚核糖体)组分与总RNA进行转录组谱分析，最终共计获得8个样本。