期刊
METHODS
卷 26, 期 1, 页码 37-47出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/S1046-2023(02)00006-3
关键词
chromatin immunoprecipitation; transcription factors; target genes; CpG microarrays; shotgun cloning; E2F
资金
- NCI NIH HHS [CA07175, CA45250, CA09681] Funding Source: Medline
- NATIONAL CANCER INSTITUTE [T32CA009681, P30CA007175] Funding Source: NIH RePORTER
The standard chromatin immunoprecipitation (ChIP) assay is used to examine the specific association of transcription factors with DNA in the context of living cells. Here we review two modifications to this protocol which are designed to identify novel target genes of transcription factors in mammalian cells. The main advantage to both of these approaches is that only DNA sequences directly bound by a factor within the context of a living cell will be identified. Therefore, artifacts associated with overexpression and/or alterations in signaling pathways arc avoided, The first modification we describe, a ChIP cloning strategy, can be used to isolate any genomic fragment specifically associated with a particular factor. It requires no special equipment or reagents other than a high-affinity antibody to be used for immunoprecipitation of the factor of interest. However, it is most useful for the isolation of a small number of genomic targets. In contrast, the second modification, which combines ChIP with specialized CpG microarrays, is ideal for a more global analysis of target genes. Advantages, common problems, and detailed protocols for these two ChIP techniques are discussed. (C) 2002 Elsevier Science (USA). All rights reserved.
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