Qubit dsdna hs assay kit

To investigate possible mechanisms for the robust decrease in physical activity in female ArKO mice, RNAseq data were collected from the NAc brain region for male and female WT and ArKO mice. The average total reads were 64,519,492.72 reads with an average of 56,945,785.76 reads after quality control (Supplementary Table 1). The average number of mapped reads was 56,501,086.96 translating to 99.214% of reads being mapped. We previously demonstrated this average number of reads to be sufficient for eukaryotic transcriptome data (Johnson et al., 2017 ). High-throughput sequencing was performed at the University of Missouri DNA Core Facility (Columbia, MO) as described previously (Ortega et al., 2019 (link)). Libraries were constructed following the manufacturer’s protocol with reagents supplied in Illimina’s TruSeq mRNA Stranded Library Preparation kit.
Briefly, the poly-A containing mRNA is purified from total RNA. RNA is fragmented and double-stranded cDNA is generated from fragmented RNA, and the index containing adapters are ligated to the ends. The final construct of each purified library was evaluated using the Fragment Analyzer automated electrophoresis system, quantified with the Qubit fluorometer using the Qubit HS dsDNA assay kit, and diluted according to Illumina’s standard sequencing protocol for sequencing on the NextSeq 500.

RNA obtained from sorted BM AML cells of two DPP4^+/+ and two DPP4^−/− AML cells transplanted mice, was purified with QIAGEN miRNeasy Mini Kit. The sample concentration was determined by Qubit fluorometer (Invitrogen, Waltham MA) using the Qubit HS RNA assay kit, and RNA integrity was assessed using the Fragment Analyzer automated electrophoresis system. Next, poly-A containing mRNA was purified from total RNA (1mg), RNA was fragmented, double-stranded cDNA was generated from fragmented RNA, and the index-containing adapters were ligated to the ends. Libraries were constructed following the manufacturer’s protocol with reagents supplied in Illumina’s TruSeq mRNA stranded sample preparation kit. The amplified cDNA constructs were purified by addition of Axyprep Mag PCR Clean-up beads. The final construct of each purified library was evaluated using the Fragment Analyzer automated electrophoresis system, quantified with the Qubit fluorometer using the Qubit HS dsDNA assay kit, and diluted according to Illumina’s standard sequencing protocol for sequencing on the NextSeq 500. High-throughput sequencing was performed at the University of Missouri DNA Core Facility.