HEK293T Mtr4-KD and Control Ribo-Seq Reads for Ribosome Profiling

<p>Ribo-seq reads from Mtr4-knockdown and siRNA control cells used for ribosome profiling under Mtr4-depleted conditions.</p> <p>Cell line: HEK293T</p> <p>Transfection: siRNA, 72 hours</p> <p>Cells were lysed with cycloheximide lysis buffer, ribosomes were isolated by Ambion RNaseI digestion of unprotected RNA followed by sucrose gradient fractionation. Footprints were obtained by TRIZOL-chloroform extraction from ribosome fractions followed by urea-PAGE gel extraction. Libraries were made using the NEBNext Small RNA Library Prep Kit and rRNA fragments were depleted with custom biotinylated oligos.</p> <p>Sequencing was performed using an Illumina NextSeq 500 for 400 million single-end reads.</p> <p>Reads were converted into FASTQ format by Illumina BaseSpace. Adapters were removed by CutAdapt, remaining rRNA reads were removed by Bowtie2 alignment vs published rRNA sequences.</p>