title: GSM8193774: MDSC, RNA_seq, Nrp1_high, rep2; Mus musculus; RNA-Seq
library_construction_protocol: For RNA-seq, a total of 8×104 Nrp1high monocytes or Nrp1low monocytes were sorted from the spleens of C57BL/6J neonatal mice (7-day-old) and lysed in 5 µl 10× lysis buffer. The SMART_x0002_Seq v4 Ultra Low Input RNA Kit (Clontech) was used to prepare the low-input library. Briefly, the lysed RNA was denatured at 72°C for 3 min, and PCR was performed in a mixture of 5× Ultra Low First Strand Buffer, SMART-Seq v4 oligonucleotide, and RNase inhibitor (42°C for 90 min, 70°C for 10 min, and 4°C for hold). PCR Primer II A amplifies cDNA from the SMART sequences introduced by 3′ SMART-Seq CDS Primer II A and the SMART-Seq v4 Oligonucleotide and then purification of amplified cDNA using the Agencourt AMPure XP Kit (Beckman Coulter).  For ATAC-seq, a total of 5×104 Nrp1high monocytes or Nrp1low monocytes were sorted from the spleens 8 of C57BL/6 neonatal mice (7-day-old) and washed once with PBS before they were resuspended by 50 l cold lysis buffer for 10 min on ice. After that, the cell nuclei were resuspended in 50 l of transposition mixture, and the mixture was incubated at 37 °C for 30 min for DNA fragmentation. The DNA fragments were purified with VAHTS DNA Clean Beads. For RNA-seq, a cDNA library constructed by technology from the pooled RNA was sequenced on the Illumina  platform using a 2×150 bp paired-end sequencing protocol.  For ATAC-seq, libraries were constructed using the TruePrep DNA Library Prep Kit V2 for Illumina (TD501, Vazyme) and the TruePrep Index Kit V2 for Illumina(TD202, Vazyme). Libraries were 2×150 bp paired-end sequenced on a HiSeq X Ten platform (Illumina) with the sequencing read length.