title: GSM1429062: E12.5 Kidney Single Cell RNA-Seq-cell 23; Mus musculus; RNA-Seq
library_construction_protocol: The single cell suspension was loaded into the Fluidigm C1 machine. The cell capture, wash and lysis were carried out according to standard Fluidigm protocols. Following capture individual capture sites were examined microscopically to record which chambers received single cells. The reverse transcription and amplification reactions were carried out using standard Fluidigm (C1 Single-Cell Auto Prep System) methods with the SMARTer Ultra Low Input RNA for Illumina Sequencing Kit (Clontech) and the Advantage 2 PCR Kit (Clontech).   The libraries were prepared with the Nextera XT DNA Sample Preparation Kit (Illumina Technologies).  1ng cDNA was suspended in Tagment DNA Buffer, and tagmentation (fragmentation and tagging with the adaptors) was performed with the Nextera enzyme (Tagment DNA Enzyme).  Tagged and fragmented cDNA was purified with DNA Clean & Concentrator-5 (Zymo Research).  The libraries were prepared by PCR with Nextera PCR Master Mix, PCR Primer Cocktail, 2 Nextera Indexes (Index 1 (i7) N701-N712 & Index 2(i5) N501-N508) and barcoded according to the following program: one cycle 98C for 30s, 5 cycles of 98C for 10s, 63C for 30s, 72C for 3min.  The purified DNA was captured on an Illumina flow cell for cluster generation.  Libraries were sequenced on the Illumina HiSeq2500 following the manufacturer's protocols.