Gene expression profiles of P7 bladder epithelium compartments isolated from Upk1b mice. (GUDMAP Series ID: 34)

The long term objective is to create an encyclopedia of the expression levels of all genes in multiple components of the developing genitourinary tract The central thesis is straightforward. The combination of microdissected tissues and FACS sorted cells plus microarray analysis offers a powerful, efficient and effective method for the creation of a global gene expression atlas of the developing urogenital system. Microarrays with essentially complete genome coverage can be used to quantitate expression levels of every gene. The ensuing rapid read-out provides an expression atlas that is more sensitive, more economical and more complete than would be possible by in situ hybridizations alone. The data submitted here delineates the gene expression profiles of the epithelial compartments of the P7 mouse bladder. At postnatal day 7 mice were euthanized by CO2 asphyxiation and the bladders were removed and cut at the bladder neck. The bladder was cut into rings and treated with 1 mg/ml trypsin in Tyrode's solution for 30 minutes at 37 °C. The layers were separated by gentle microdissection and the epithelial layer was further digested in a mixture of Blendzymes 1 & 4 at 37 °C until a single cell suspension was obtained. The samples were placed in RLT and stored at -80 °C.RNA was prepared and the Epicentre 2-round amplification scheme described under the Lessard Group Protocols on the GUDMAP pages were performed. The Affymetrix GeneChip Mouse Genome 430 2.0 Array was used to interrogate the amplified RNA.本研究的长期目标为构建发育中泌尿生殖道多组分的全基因表达水平百科全书。其核心论点清晰明确：将显微切割组织、荧光激活细胞分选（FACS）细胞与微阵列分析相结合，可为发育中的泌尿生殖系统构建全局基因表达图谱提供高效且功能强大的技术手段。具备全基因组覆盖度的微阵列可用于定量所有基因的表达水平，由此获得的快速检测结果可生成比单纯原位杂交更灵敏、更经济且更完整的表达图谱。本次提交的数据描绘了出生后第7天（P7）小鼠膀胱上皮组分的基因表达谱。实验过程如下：于出生后第7天通过二氧化碳窒息法处死小鼠，取出膀胱并在膀胱颈处切开。将膀胱切成环状组织，置于含1mg/ml胰蛋白酶的台氏液（Tyrode's solution）中，于37℃条件下孵育30分钟。通过轻柔显微切割分离各组织层，随后将上皮层置于Blendzymes 1与4的混合酶液中，于37℃继续消化直至获得单细胞悬液。将样本置于RLT裂解液中并于-80℃保存。随后完成RNA制备，并按照GUDMAP平台上Lessard课题组方案中所述的Epicentre两轮扩增流程进行操作。采用Affymetrix GeneChip小鼠基因组430 2.0阵列对扩增后的RNA进行检测分析。