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Cell type–specific mRNA purification by translating ribosome affinity purification (TRAP)
Open accessAbstract
Cellular diversity and architectural complexity create barriers to understanding the function of the mammalian CNS at a molecular level. To address this problem, we have recently developed a methodology that provides the ability to profile the entire translated mRNA complement of any genetically defined cell population. This methodology, which we termed translating ribosome affinity purification, or TRAP, combines cell type-specific transgene expression with affinity purification of translating ribosomes. TRAP can be used to study the cell type-specific mRNA profiles of any genetically defined cell type, and it has been used in organisms ranging from Drosophila melanogaster to mice and human cultured cells. Unlike other methodologies that rely on microdissection, cell panning or cell sorting, the TRAP methodology bypasses the need for tissue fixation or single-cell suspensions (and the potential artifacts that these treatments introduce) and reports on mRNAs in the entire cell body. This protocol provides a step-by-step guide to implement the TRAP methodology, which takes 2 d to complete once all materials are in hand.
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“The TRAP approach91,92 relies on the genetic labelling of the ribosomal protein L10a with the enhanced green fluorescent protein (eGFP) using Eef1a1LSL.eGFPL10a/+ mice60.”
From Lymphoid gene expression supports neuroprotective microglia function (2025) · cites this paper as [92]
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Cites
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“RNAwas collected from hippocampi, striata, and cortex of Aldh1l1-eGFP-L10a mice based on published protocols (21, 22).”
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“(legend continued on next page) 4 Cell 163, 1–13, October 8, 2015 ª2015 Elsevier Inc. (TRAP), which was developed to identify cell type-specific transcripts in tissues containing heterogeneous cell populations (Heiman et al., 2014).”
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Ribosome profiling reveals the what, when, where and how of protein synthesis
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“…adds a tag to a ribosomal protein of the large ribosomal subunit in a cell type–specific manner by using either bac transgenics or the Cre-lox system in mice (Doyle et al. 2008; Heiman et al. 2014; Hupe et al. 2014), but can be applied to worms, flies, plants, and yeast (King and Gerber 2016).”
Method
Metadata
- Authors
- Heiman, M., Kulicke, R., Fenster, R. J., Greengard, P., & Heintz, N.
- Affiliations
- Broad Institute (US)
- Rockefeller University (US)
- Howard Hughes Medical Institute (US)
- Corresponding author
- Heiman, Myriam
- Venue
- Nature Protocols
- Published
- 2014-05-08
- Volume
- 9
- Issue
- 6
- Pages
- 1282-1291
- ISSN
- 1750-2799 (ISSN-L)
- 1754-2189
- PMC
- PMC4102313
- OpenAlex
- W1993981485
- Cited by
- 498
- References
- 39
- Citations by year
- 2026: 36 · 2025: 49 · 2024: 54 · 2023: 52 · 2022: 54 · 2021: 44 · 2020: 49 · 2019: 48 · 2018: 45 · 2017: 30 · 2016: 21 · 2015: 12 · 2014: 4
- FWCI
- 9.16
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Nature Protocols (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc-sa)
- PMC
- Funders
- Howard Hughes Medical Institute
- National Institute of Mental Health (P50MH090963, MH090963)
- National Institute of General Medical Sciences (T32GM007739)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
- Keywords
- translating ribosome affinity purification, central nervous system, cell type-specific gene expression
- MeSH terms
- Animals, Cells, Cultured, Humans, Polyribosomes, Ribosomes, RNA, Messenger, Gene Expression Profiling, Mice, Single-Cell Analysis
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