Next-generation sequencing measuring ribosome mRNA occupancy and translation efficiency in CD4+ T cells isolated from Mettl3 KO and WT mice

50 million CD4+ naive T cells were isolated from Mettl3 KO and WT mice by StemCell CD4+ T cell isolation kits. The ribosome protected fragments and inputs were prepared strictly following the manual of Illumina TruSeq Ribo Profile (Mammalian) Kit. The libraries were generated using the same kit and subjected to sequencing. Raw sequencing reads were aligned to the mouse genome (mm10) with Tophat, and anlayzed by Tuxedo suite & HTseq. Ribosome protected fragments were isolated following Illumina TruSeq Ribo Profile Kit with CD4+ T cells from Mettl3 KO and WT mouse, and subjected to deep sequencing, in duplicate, using Illumina HiSeq 2000.