Common cell lysis procedures distort ribosome profiling analyses of gene expression

IF 10.1 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Genome Biology Pub Date : 2025-08-11 DOI:10.1186/s13059-025-03651-1

Aoife O’Connell, Alla D. Fedorova, Patrick B. F. O’Connor, Alexander V. Zhdanov, Pavel V. Baranov, Gary Loughran, Dmitry E. Andreev

引用次数: 0

Abstract

Ribosome profiling is a powerful technique used to study gene expression on a transcriptome-wide scale. It involves sequencing of mRNA fragments protected by ribosomes from ribonuclease digestion. The initial steps commonly involve cell lysis followed by centrifugation and ribonuclease digestion. We find that centrifugation depletes 329 translated mRNAs in HEK293T cells. Many of these mRNAs encode cytoskeleton proteins. This suggests that the expression of a subset of mRNAs may be significantly underestimated in most ribosome profiling experiments. We show that omitting the centrifugation step after cell lysis can resolve this issue.

查看原文本刊更多论文

普通的细胞裂解程序扭曲了基因表达的核糖体谱分析

核糖体谱分析是一种用于研究转录组范围内基因表达的强大技术。它涉及对受核糖体保护的mRNA片段进行测序，使其免受核糖核酸酶的酶切。最初的步骤通常包括细胞裂解，然后是离心和核糖核酸酶消化。我们发现，在HEK293T细胞中，离心会耗尽329种翻译mrna。许多mrna编码细胞骨架蛋白。这表明，在大多数核糖体分析实验中，mrna亚群的表达可能被严重低估。我们表明，在细胞裂解后省略离心步骤可以解决这个问题。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

求助全文

约1分钟内获得全文求助全文

来源期刊

Genome Biology Biochemistry, Genetics and Molecular Biology-Genetics

CiteScore

21.00

自引率

3.30%

发文量

241

审稿时长

2 months

期刊介绍： Genome Biology stands as a premier platform for exceptional research across all domains of biology and biomedicine, explored through a genomic and post-genomic lens. With an impressive impact factor of 12.3 (2022),* the journal secures its position as the 3rd-ranked research journal in the Genetics and Heredity category and the 2nd-ranked research journal in the Biotechnology and Applied Microbiology category by Thomson Reuters. Notably, Genome Biology holds the distinction of being the highest-ranked open-access journal in this category. Our dedicated team of highly trained in-house Editors collaborates closely with our esteemed Editorial Board of international experts, ensuring the journal remains on the forefront of scientific advances and community standards. Regular engagement with researchers at conferences and institute visits underscores our commitment to staying abreast of the latest developments in the field.