دورية أكاديمية
Optimization of Ribosome Footprinting Conditions for Ribo-Seq in Human and Drosophila melanogaster Tissue Culture Cells.
| العنوان: | Optimization of Ribosome Footprinting Conditions for Ribo-Seq in Human and Drosophila melanogaster Tissue Culture Cells. |
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| المؤلفون: | Douka K; School of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom.; LeedsOmics, University of Leeds, Leeds, United Kingdom., Agapiou M; School of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom.; LeedsOmics, University of Leeds, Leeds, United Kingdom., Birds I; School of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom.; LeedsOmics, University of Leeds, Leeds, United Kingdom., Aspden JL; School of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom.; LeedsOmics, University of Leeds, Leeds, United Kingdom. |
| المصدر: | Frontiers in molecular biosciences [Front Mol Biosci] 2022 Jan 25; Vol. 8, pp. 791455. Date of Electronic Publication: 2022 Jan 25 (Print Publication: 2021). |
| نوع المنشور: | Journal Article |
| اللغة: | English |
| بيانات الدورية: | Publisher: Frontiers Media S.A Country of Publication: Switzerland NLM ID: 101653173 Publication Model: eCollection Cited Medium: Print ISSN: 2296-889X (Print) Linking ISSN: 2296889X NLM ISO Abbreviation: Front Mol Biosci Subsets: PubMed not MEDLINE |
| أسماء مطبوعة: | Original Publication: Lausanne : Frontiers Media S.A., [2014]- |
| مستخلص: | 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 generated different footprints sizes and framing patterns in human and D. melanogaster cells. We also found that treatment of S2 cells with cycloheximide prior to footprinting impacted the distribution of footprints across ORFs, without affecting overall read length distribution and framing pattern, as previously found in other organisms. Together our results indicate that a variety of factors affect ribosome footprint quality and the nature of precise footprinting varies across species. Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. (Copyright © 2022 Douka, Agapiou, Birds and Aspden.) |
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| معلومات مُعتمدة: | MR/N000471/1 United Kingdom MRC_ Medical Research Council |
| فهرسة مساهمة: | Keywords: ORF; Ribo-seq; footprinting; mRNA translation; ribosome profiling |
| تواريخ الأحداث: | Date Created: 20220211 Latest Revision: 20220310 |
| رمز التحديث: | 20231215 |
| مُعرف محوري في PubMed: | PMC8822167 |
| DOI: | 10.3389/fmolb.2021.791455 |
| PMID: | 35145996 |
| قاعدة البيانات: | MEDLINE |
Full Text Finder | تدمد: | 2296-889X |
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| DOI: | 10.3389/fmolb.2021.791455 |