title: GSM9075204: WTKL2; Mus musculus; RNA-Seq
library_construction_protocol: The Cd11b+Ly6G-CD64+Siglec-Flow population was isolated based FACS. The cells were homogenized in 1 ml TRIzol (invitrogen). Total RNAs were prepared and the quality of RNAs was determined by agarose gel electrophoresis and spectrophotometer analysis. Poly(A) mRNA was subsequently purified from 10μg total RNA using NEBNext Oligo d(T)25 Magnetic Beads Isolation Module. First-strand complementary DNA was synthesized with NEBNext RNA First-Strand Synthesis Module. NEBNext Ultra II Non-Directional RNA Second Strand Synthesis Module was used for the synthesis of the complementary strand of first-strand cDNA. The resulting double-stranded DNA was purified and Vazyme TruePrep DNA Library Prep kit V2 was used to prepare libraries followed by sequencing on an Illumina Hiseq X Ten platform with 150-bp paired-end reads strategy (Novogene) After generation of GEMs, reverse transcription reactions were engaged barcoded full-length cDNA followed by the disruption of emulsions using the recovery agent and cDNA clean up with DynaBeads Myone Silane Beads (Thermo Fisher Scientific). The cDNAs were then amplified by PCR with 12 cycles. Subsequently, the amplified cDNA was fragmented, end-repaired, A-tailed, index adaptor ligated and library amplification via PCR (8 cycles). The purified libraries were sequenced on the Illumina NovaSeq 6000 with 28 cycles of read1, 8 cycles of i7 index, and 91 cycles of read2. And finally, 150 bp paired-end reads were generated.