The brain endothelial cell translatome analysis in the visual cortex at plastic and nonplastic ages

In this study, we provide the translatome data in the brain endothelial cells (BECs) of the visual cortex at 3 different time points (pre-critical period (P10), critical period (P30), post-critical period (P100)) by using translating ribosome affinity purification (TRAP) combined with high-throughput RNA sequencing analysis in Tie2-Cre:Ribo-Tag mice, which are designed for expressing hemagglutinin A (HA)-tagged RPL22, a ribosomal protein in endothelial cells depending on CRE recombinase. To obtain a high-quality BEC pool, we modified the TRAP method by adding the capillary isolation process before the immunoprecipitation step and finally verified BEC-specific translating mRNA purification using RT-PCR.本研究中，我们利用Tie2-Cre:Ribo-Tag小鼠——该工程小鼠可依赖CRE重组酶（CRE recombinase）在内皮细胞中表达血凝素A（HA）标记的核糖体蛋白RPL22——结合翻译核糖体亲和纯化（translating ribosome affinity purification, TRAP）与高通量RNA测序分析，获取了视觉皮层脑内皮细胞（brain endothelial cells, BECs）在3个关键时间节点的翻译组数据，分别为关键期前（P10）、关键期（P30）及关键期后（P100）。为获得高质量的脑内皮细胞样本池，我们对TRAP实验方法进行了优化，在免疫沉淀步骤前新增毛细血管分离流程，并最终通过逆转录聚合酶链式反应（RT-PCR）验证了脑内皮细胞特异性翻译mRNA的纯化效果。