title: RNA seq of MEF : Amcf1 Knockout under Starvation condition Replicate 2
design: Total RNA was extracted using the RNeasy Mini Kit (Qiagen, 74104) with on-column DNase digestion (RNase-Free DNase Set; Qiagen, 79254) according to the manufacturers protocol. Samples with RNA integrity number (RIN) > 7 were used for library preparation. Poly(A)+ RNA was isolated using the NEBNext Ploy(A) mRNA Magnetic Isolation Module (New England Biolabs, E7490L) and libraries were constructed using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs, E7760L) along with NEBNext Multiplex Oligos for Illumina (New England Biolabs, E6609S), following the manufacturers instructions. Library quality was assessed using the Agilent High Sensitive DNA Kit (Agilent, 5067-4626) on a Bioanalyzer 2100 (Agilent Technologies, Santa Clara, USA), and concentrations were quantified by qPCR using Collibri Library Quantification kit (Thermo Fisher Scientific, A38524500). Equimolar libraries were pooled and subjected to paired-end 150-cycle sequencing on an Illumina NovaSeq platform (Illumina, Inc., USA)