RNA-Seq of Mus musculus: Mixed tissue from heart, kidney, liver, muscle, spleen and testis
Total RNA was extracted by grinding tissue in TRIzol reagent (TIANGEN) on dry ice and processed following the protocol provided by the manufacturer.The poly-A RNAs were enriched from total RNA using Dynabeads mRNA Purification Kit (Cat#61006, Invitrogen) and fragmented into small pieces using fragmentation reagent in MGIEasy RNA Library Prep Kit V3.1 (Cat# 1000005276, MGI). The first strand cDNA was synthesized using random primes and reverse transcriptase, followed by second strand cDNA synthesis. The synthesized cDNA was end-repaired, A-tailing added and ligated to the sequencing adapters according to library construction protocol. The cDNA fragments were amplified by PCR and purified with MGIEasy DNA Clean beads (CAT#1000005279, MGI). Library was analyzed on the Agilent Technologies 2100 bioanalyzer. The double stranded PCR products were heat denatured and circularized by the splint oligo sequence in MGIEasy Circularization ModuleCAT#1000005260, MGI. The single strand circle DNA (ssCir DNA) were formatted as the final library. The qualified libraries were sequenced on DNBSEQ-T7RS platform.