Abstract
Half of all human transcription factors are zinc finger proteins and yet very little is known concerning the biological role of the majority of these factors. In particular, very few genome-wide studies of the in vivo binding of zinc finger factors have been performed. Based on in vitro studies and other methods that allow selection of high affinity-binding sites in artificial conditions, a zinc finger code has been developed that can be used to compose a putative recognition motif for a particular zinc finger factor (ZNF). Theoretically, a simple bioinformatics analysis could then predict the genomic locations of all the binding sites for that ZNF. However, it is unlikely that all of the sequences in the human genome having a good match to a predicted motif are in fact occupied in vivo (due to negative influences from repressive chromatin, nucleosomal positioning, overlap of binding sites with other factors, etc). A powerful method to identify in vivo binding sites for transcription factors on a genome-wide scale is the chromatin immunoprecipitation (ChIP) assay, followed by hybridization of the precipitated DNA to microarrays (ChIP-chip) or by high throughput DNA sequencing of the sample (ChIP-seq). Such comprehensive in vivo binding studies would not only identify target genes of a particular zinc finger factor, but also provide binding motif data that could be used to test the validity of the zinc finger code. This chapter describes in detail the steps needed to prepare ChIP samples and libraries for high throughput sequencing using the Illumina GA2 platform and includes descriptions of quality control steps necessary to ensure a successful ChIP-seq experiment.
| Original language | English (US) |
|---|---|
| Title of host publication | Methods in Molecular Biology |
| Pages | 437-455 |
| Number of pages | 19 |
| Volume | 649 |
| DOIs | |
| State | Published - 2010 |
Publication series
| Name | Methods in Molecular Biology |
|---|---|
| Volume | 649 |
| ISSN (Print) | 10643745 |
Fingerprint
Keywords
- ChIP-seq
- chromatin immunoprecipitation
- next generation sequencing
- Zinc fingers
ASJC Scopus subject areas
- Molecular Biology
- Genetics
- Medicine(all)
Cite this
Using ChIP-seq technology to identify targets of zinc finger transcription factors. / O'Geen, Henriette; Frietze, Seth; Farnham, Peggy J.
Methods in Molecular Biology. Vol. 649 2010. p. 437-455 (Methods in Molecular Biology; Vol. 649).Research output: Chapter in Book/Report/Conference proceeding › Chapter
}
TY - CHAP
T1 - Using ChIP-seq technology to identify targets of zinc finger transcription factors
AU - O'Geen, Henriette
AU - Frietze, Seth
AU - Farnham, Peggy J.
PY - 2010
Y1 - 2010
N2 - Half of all human transcription factors are zinc finger proteins and yet very little is known concerning the biological role of the majority of these factors. In particular, very few genome-wide studies of the in vivo binding of zinc finger factors have been performed. Based on in vitro studies and other methods that allow selection of high affinity-binding sites in artificial conditions, a zinc finger code has been developed that can be used to compose a putative recognition motif for a particular zinc finger factor (ZNF). Theoretically, a simple bioinformatics analysis could then predict the genomic locations of all the binding sites for that ZNF. However, it is unlikely that all of the sequences in the human genome having a good match to a predicted motif are in fact occupied in vivo (due to negative influences from repressive chromatin, nucleosomal positioning, overlap of binding sites with other factors, etc). A powerful method to identify in vivo binding sites for transcription factors on a genome-wide scale is the chromatin immunoprecipitation (ChIP) assay, followed by hybridization of the precipitated DNA to microarrays (ChIP-chip) or by high throughput DNA sequencing of the sample (ChIP-seq). Such comprehensive in vivo binding studies would not only identify target genes of a particular zinc finger factor, but also provide binding motif data that could be used to test the validity of the zinc finger code. This chapter describes in detail the steps needed to prepare ChIP samples and libraries for high throughput sequencing using the Illumina GA2 platform and includes descriptions of quality control steps necessary to ensure a successful ChIP-seq experiment.
AB - Half of all human transcription factors are zinc finger proteins and yet very little is known concerning the biological role of the majority of these factors. In particular, very few genome-wide studies of the in vivo binding of zinc finger factors have been performed. Based on in vitro studies and other methods that allow selection of high affinity-binding sites in artificial conditions, a zinc finger code has been developed that can be used to compose a putative recognition motif for a particular zinc finger factor (ZNF). Theoretically, a simple bioinformatics analysis could then predict the genomic locations of all the binding sites for that ZNF. However, it is unlikely that all of the sequences in the human genome having a good match to a predicted motif are in fact occupied in vivo (due to negative influences from repressive chromatin, nucleosomal positioning, overlap of binding sites with other factors, etc). A powerful method to identify in vivo binding sites for transcription factors on a genome-wide scale is the chromatin immunoprecipitation (ChIP) assay, followed by hybridization of the precipitated DNA to microarrays (ChIP-chip) or by high throughput DNA sequencing of the sample (ChIP-seq). Such comprehensive in vivo binding studies would not only identify target genes of a particular zinc finger factor, but also provide binding motif data that could be used to test the validity of the zinc finger code. This chapter describes in detail the steps needed to prepare ChIP samples and libraries for high throughput sequencing using the Illumina GA2 platform and includes descriptions of quality control steps necessary to ensure a successful ChIP-seq experiment.
KW - ChIP-seq
KW - chromatin immunoprecipitation
KW - next generation sequencing
KW - Zinc fingers
UR - http://www.scopus.com/inward/record.url?scp=77957961816&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=77957961816&partnerID=8YFLogxK
U2 - 10.1007/978-1-60761-753-2_27
DO - 10.1007/978-1-60761-753-2_27
M3 - Chapter
C2 - 20680851
AN - SCOPUS:77957961816
SN - 9781607617525
VL - 649
T3 - Methods in Molecular Biology
SP - 437
EP - 455
BT - Methods in Molecular Biology
ER -