Dual randomization of oligonucleotides to reduce the bias in ribosome-profiling libraries.

ABSTRACT: Protein translation is at the heart of cellular metabolism and its in-depth characterization is key for many lines of research. Recently, ribosome profiling became the state-of-the-art method to quantitatively characterize translation dynamics at a transcriptome-wide level. However, the strategy of library generation affects its outcomes. Here, we present a modified ribosome-profiling protocol starting from yeast, human cells and vertebrate brain tissue. We use a DNA linker carrying four randomized positions at its 5' end and a reverse-transcription (RT) primer with three randomized positions to reduce artifacts during library preparation. The use of seven randomized nucleotides allows to efficiently detect library-generation artifacts. We find that the effect of polymerase chain reaction

SUBMITTER: Lecanda A 

PROVIDER: S-EPMC5024760 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature