RNA-sequencing experiments comparing P0 control and RXRG/RARb double mutants frontal cortex

To understand the functional significance of RA signaling through the RXRG/RARB heterodimer in the developing cortex, we performed RNA-seq experiments comparing different regions of the P0 mouse frontal cortex (mPFC, secondary motor cortex (MOs)/ primary motor cortex (MOp), and OFC ) in Rarb and Rxrg dKO and WT littermate controls. Freshly dissected P0 brains were frozen at -80C, and moved to RNALater 12-16 hrs prior to further dissection. Medial frontal, motor, and orbitofrontal regions were microdissected. RNA was isolated using Rneasy plus micro kit (Qiagen), and cDNA libraries were created using SMART-seq v4 Ultra Low Input Kit (Takara) and Nextera XT DNA library Prep Kit (Illumina). Libraries were normalized and sequenced at the Yale Center for Genomic Analysis (YGCA) using the NovaSeq with 100 bp paired end reads.为阐明通过RXRG/RARB异二聚体介导的视黄酸（retinoic acid, RA）信号通路在发育中的大脑皮层的功能意义，我们开展了RNA测序（RNA-seq）实验，对比了出生后0天（P0）小鼠前额叶皮层的不同区域：内侧前额叶皮层（medial prefrontal cortex, mPFC）、次级运动皮层（secondary motor cortex, MOs）/初级运动皮层（primary motor cortex, MOp）以及眶额皮层（orbitofrontal cortex, OFC），实验分组为Rarb与Rxrg双基因敲除（double knockout, dKO）小鼠及其野生型（wild type, WT）同窝对照。 我们将新鲜解剖获取的P0小鼠脑组织置于-80℃冷冻保存，并在后续解剖前12~16小时转移至RNALater保存液中。随后对内侧前额叶、运动皮层及眶额叶区域进行显微解剖。总RNA提取采用Qiagen的Rneasy plus micro试剂盒，cDNA文库构建分别使用Takara的SMART-seq v4 Ultra Low Input Kit与Illumina的Nextera XT DNA文库制备试剂盒。 文库经归一化处理后，由耶鲁大学基因组分析中心（Yale Center for Genomic Analysis, YGCA）使用NovaSeq测序平台完成100 bp双端测序。