TY - CHAP
T1 - LncRNA pulldown combined with mass spectrometry to identify the novel LncRNA-associated proteins
AU - Xing, Zhen
AU - Lin, Chunru
AU - Yang, Liuqing
N1 - Publisher Copyright:
© Springer Science+Business Media New York 2016.
PY - 2016/1/1
Y1 - 2016/1/1
N2 - Long noncoding RNAs (LncRNAs) are nonprotein-coding transcripts longer than 200 nucleotides in length. The recent studies have revealed that at least nearly 80 % transcripts in human cells are lncRNA species. Based on their genomic location, most lncRNAs can be characterized as large intergenic noncoding RNAs, natural antisense transcripts, pseudogenes, long intronic ncRNAs, as well as other divergent transcripts. However, despite mounting evidences suggesting that many lncRNAs are likely to be functional, only a small proportion has been demonstrated to be biologically and physiologically relevant due to their lower expression levels and current technique limitations. Thus, there is a greater need to design and develop new assays to investigate the real function of lncRNAs in depth in various systems. Indeed, several methods such as genome-wide chromatin immunoprecipitation-sequencing (ChIP-seq), RNA immunoprecipitation followed by sequencing (RIP-seq) have been developed to examine the genome localization of lncRNAs and their interacting proteins in cells. Here we describe an open-ended method, LncRNA pulldown assay, which has been frequently used to identify its interacting protein partners in the cellular context. Here we provide a detailed protocol for this assay with hands-on tips based on our own experience in working in the lncRNA fields.
AB - Long noncoding RNAs (LncRNAs) are nonprotein-coding transcripts longer than 200 nucleotides in length. The recent studies have revealed that at least nearly 80 % transcripts in human cells are lncRNA species. Based on their genomic location, most lncRNAs can be characterized as large intergenic noncoding RNAs, natural antisense transcripts, pseudogenes, long intronic ncRNAs, as well as other divergent transcripts. However, despite mounting evidences suggesting that many lncRNAs are likely to be functional, only a small proportion has been demonstrated to be biologically and physiologically relevant due to their lower expression levels and current technique limitations. Thus, there is a greater need to design and develop new assays to investigate the real function of lncRNAs in depth in various systems. Indeed, several methods such as genome-wide chromatin immunoprecipitation-sequencing (ChIP-seq), RNA immunoprecipitation followed by sequencing (RIP-seq) have been developed to examine the genome localization of lncRNAs and their interacting proteins in cells. Here we describe an open-ended method, LncRNA pulldown assay, which has been frequently used to identify its interacting protein partners in the cellular context. Here we provide a detailed protocol for this assay with hands-on tips based on our own experience in working in the lncRNA fields.
KW - Long noncoding RNAs
KW - Mass spectrometry
KW - RNA-protein interaction
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U2 - 10.1007/978-1-4939-3378-5_1
DO - 10.1007/978-1-4939-3378-5_1
M3 - Chapter
C2 - 26721478
AN - SCOPUS:84953259172
T3 - Methods in Molecular Biology
SP - 1
EP - 9
BT - Methods in Molecular Biology
PB - Humana Press Inc.
ER -