RNA-seq of the E14.5 lacrimal gland

The goal of this study is to find the transcriptional targets downstream of PI3K signaling during lacrimal gland development. We performed laser capture microdissection of the lacrimal gland epithelial tissue at embryonic day E14.5 from control embryos and mutant embryos containing lacrimal gland-specific deletion of PI3K subunits. After tissue harvest, RNA was extracted, conversion to cDNA and amplification was performed by Clontech SMART-seq v4 Ultra low input RNA kit,and cDNA library construction was performed using Nextera XT DNA library preparation kit by core facility at Columbia university prior to RNA sequencing. After preparation of cDNA library, each sample was sequenced using Illumina platform For this experiment, 3 control samples and 6 mutant samples were analyzed.本研究旨在探究泪腺发育过程中磷脂酰肌醇3-激酶（PI3K）信号通路下游的转录靶标。我们分别选取对照组胚胎与泪腺特异性敲除PI3K亚基的突变体胚胎，在胚胎发育第14.5天（E14.5）对其泪腺上皮组织开展激光捕获显微切割操作。组织收集完成后，提取总RNA，随后采用Clontech SMART-seq v4 超低起始量RNA试剂盒完成cDNA合成与扩增；再由哥伦比亚大学核心实验设施采用Nextera XT DNA文库制备试剂盒构建cDNA文库，随后进行RNA测序。cDNA文库制备完成后，所有样本均采用Illumina测序平台完成测序。本实验共分析3个对照样本与6个突变体样本。