Illumina HiSeq 2000 paired end sequencing; GSM1972076: single cell RNA-seq [NPC1_S1]; Mus musculus; RNA-Seq

single cell RNA-seq [NPC1_S1]

The extent of transcriptional diversity in mouse NPCs is likely to be influenced by a variety of unexamined factors that include programmed cell death, genomic mosaicism as well as a variety of “environmental” influences such as changes in exposure to signaling lipids. We therefore used scRNA-seq to assess a cohort of cortical NPCs from an embryonic mouse. We demonstrate that PAGODA (Pathway And Geneset OverDispersion Analysis) effectively recovers the known neuroanatomical and functional organization of NPCs, identifying multiple aspects of transcriptional heterogeneity within the developing mouse cortex that are difficult to discern by the existing heterogeneity analysis approaches. Overall design: Examination of mouse NPC transcriptional heterogeneity via single cell RNA-seq

Characterizing transcriptional heterogeneity through pathway and gene set overdispersion analysis (PAGODA)

Submitted by Gene Expression Omnibus on 19-JAN-2016

Timed-pregnant mice were sacrificed by deep anesthesia followed by cervical dislocation. The embryos were quickly removed and cortical hemispheres were isolated, ganglionic eminences removed, and all pups brains were pooled. Single cells were isolated by gentle trituration in ice-cold phosphate buffered saline containing 2 mM EGTA (PBSE) using P1000 tips with decreasing bore diameter. Cells were then filtered through a 40 uM nylon cell strainer and stained with propidium iodide (PI), a live-dead stain, and fluorescence activated single cell sorting (FACS) was performed selecting for PI negative cells. Samples remained on ice throughout the process and total processing time from cervical dislocation to sorting was limited to 2 hours. Single cells were sorted directly into cell lysis buffer provided in the Clontech SMARTer® Ultra™ Low RNA Kit for Illumina® Sequencing (cat # 634936), and sequencing libraries were generated using the manufacturer’s protocol. Resulting libraries were sequenced on the Illumina® HiSeqTM 2000 sequencing platform.

Characterizing transcriptional heterogeneity through pathway and gene set overdispersion analysis (PAGODA)

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