Optimization of Ribosome Footprinting Conditions for Ribo-Seq in Human and <i>Drosophila melanogaster</i> Tissue Culture Cells.

ABSTRACT: Our understanding of mRNA translation and its regulation has been transformed by the development of ribosome profiling. This approach relies upon RNase footprinting of translating ribosomes in a precise manner to generate an accurate snapshot of ribosome positions with nucleotide resolution. Here we tested a variety of conditions, which contribute to the preciseness of ribosome footprinting and therefore the success of ribosome profiling. We found that NaCl concentration, RNaseI source, RNaseI amount, and temperature of footprinting all contributed to the quality of ribosome footprinting in human neuroblastoma SH-SY5Y cells. These ideal conditions for footprinting also improved footprint quality when used with Drosophila melanogaster S2 cells. Footprinting under the same conditions

SUBMITTER: Douka K 

PROVIDER: S-EPMC8822167 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature