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Method Article
Yuanchao Xue
Key Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0002-8113-2333
License:
This work is licensed under a CC BY 4.0 License. Read Full License
RNA-binding proteins (RBPs) directly interact with various RNAs in living cells to regulate their processing, translation, and stability. Identifying the precise binding sites of RBPs is critical for appreciating their physiological or pathological roles in germline and early embryo development. Current methods typically need millions of cells to map RBP binding positions, which prevents us from appreciating the crucial role of RBPs in early development. Here, we present the LACE-seq method for unbiased mapping of RBP-binding sites at single-nucleotide resolution in fewer cells or even single oocytes. LACE-seq depends on RBP-mediated reverse transcription termination, and linear amplification of the cDNA ends for deep sequencing. To further promote its application, we describe a step-by-step protocol about how to construct a successful LACE-seq library.
Keywords
RNA-binding protein; deep sequencing; binding site
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Associated Publications
Global profiling of RNA-binding protein target sites by LACE-seq
by Ruibao Su, Li-Hua Fan, Changchang Cao, +16
Nature Cell Biology (09 June, 2021)
An open repository of community-contributed protocols sponsored by Nature Research.
Learn more about Protocol Exchange
Method Article
Yuanchao Xue
Key Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0002-8113-2333
Version History
CURRENT STATUS: Posted
Version 1
Posted 10 Jun, 2021
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
RNA-binding proteins (RBPs) directly interact with various RNAs in living cells to regulate their processing, translation, and stability. Identifying the precise binding sites of RBPs is critical for appreciating their physiological or pathological roles in germline and early embryo development. Current methods typically need millions of cells to map RBP binding positions, which prevents us from appreciating the crucial role of RBPs in early development. Here, we present the LACE-seq method for unbiased mapping of RBP-binding sites at single-nucleotide resolution in fewer cells or even single oocytes. LACE-seq depends on RBP-mediated reverse transcription termination, and linear amplification of the cDNA ends for deep sequencing. To further promote its application, we describe a step-by-step protocol about how to construct a successful LACE-seq library.
Loading...
Associated Publications
Global profiling of RNA-binding protein target sites by LACE-seq
by Ruibao Su, Li-Hua Fan, Changchang Cao, +16
Nature Cell Biology (09 June, 2021)