GSM4405433: Ngn3Cre_P12_2; Mus musculus; RNA-Seq
Library preparation was conducted following the modified STRT-seq protocol (Dong et al., 2018, Genome biology). Briefly, after cell sorting, single Ins1-RFP+ or Ngn3-Cre; Rosa-RFP+ cells were transferred quickly into lysis buffer by mouth pipette. Next, we reverse-transcribed the RNA and amplified the resulting cDNA over 18 cycles of PCR. The quality of the cDNA was assessed by qPCR of the housekeeping gene Gapdh. cDNA of single cells with good quality was pooled together and purified. Biotinylated pre-indexed primers were added to the cDNA during an additional 4 cycles of PCR. Approximately 300 ng of purified PCR product was sheared to approximately 300 bp by sonication. Sequencing libraries were prepared using the Kapa Hyper Prep Kit (KK8502, Kapa Biosystems).