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Genome-scale identification of Caenorhabditis elegans regulatory elements by tiling-array mapping of DNase I hypersensitive sites.

Shi B, Guo X, Wu T, Sheng S, Wang J, Skogerbø G, Zhu X, Chen R - BMC Genomics (2009)

Bottom Line: Many DHSs appeared to be associated with annotated non-coding RNAs and recently detected transcripts of unknown function.It has been reported that nematode highly conserved non-coding elements were associated with cis-regulatory elements, and we also found that DHSs, particularly distal intergenic DHSs, were significantly enriched in regions that were conserved between the C. elegans and C. briggsae genomes.We describe the first genome-wide analysis of C. elegans DHSs, and show that the distribution of DHSs is strongly associated with functional elements in the genome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bioinformatics Laboratory and National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, PR China. shibaochen@moon.ibp.ac.cn

ABSTRACT

Background: A major goal of post-genomics research is the integrated analysis of genes, regulatory elements and the chromatin architecture on a genome-wide scale. Mapping DNase I hypersensitive sites within the nuclear chromatin is a powerful and well-established method of identifying regulatory element candidates.

Results: Here, we report the first genome-wide analysis of DNase I hypersensitive sites (DHSs) in Caenorhabditis elegans. The data was obtained by hybridizing DNase I-treated and end-captured material from young adult worms to a high-resolution tiling microarray. The data show that C. elegans DHSs were significantly enriched within intergenic regions located 2 kb upstream and downstream of coding genes, and also that a considerable fraction of all DHSs mapped to intergenic positions distant to annotated coding genes. Annotated transcribed loci were generally depleted in DHSs relative to intergenic regions, but DHSs were nonetheless enriched in coding exons and UTRs, whereas introns were significantly depleted in DHSs. Many DHSs appeared to be associated with annotated non-coding RNAs and recently detected transcripts of unknown function. It has been reported that nematode highly conserved non-coding elements were associated with cis-regulatory elements, and we also found that DHSs, particularly distal intergenic DHSs, were significantly enriched in regions that were conserved between the C. elegans and C. briggsae genomes.

Conclusion: We describe the first genome-wide analysis of C. elegans DHSs, and show that the distribution of DHSs is strongly associated with functional elements in the genome.

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Protocol outline for the genome-scale mapping of C. elegans DHSs by tiling microarray analysis.
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Figure 1: Protocol outline for the genome-scale mapping of C. elegans DHSs by tiling microarray analysis.

Mentions: The protocol for the genome-scale mapping of C. elegans DNase I hypersensitive sites (DHSs) by the tiling microarray is summarized in Figure 1. The Affymetrix C. elegans Tiling 1.0R array contains ~3.2 million 25-mer oligonucleotide probe pairs covering the Watson strand of the entire non-repetitive genome at an average resolution of 25 bp. Synchronized worms in the young adult (YA) stage were treated with floxuridine (FUdR) for more than 8 hours to reduce the background signal from reproduction, without any apparent effect on vitality and longevity [6,7]. Extracted nuclei were digested with different concentrations of DNase I (Figure 2), and samples treated with 240 and 480 U/ml (along with DNA from untreated nuclei) were applied to the tiling assays. The entire procedure was replicated after an interval of about one month, and quantile-normalization was performed on the biological replicates within treatment and control groups [10]. To identify probes that are significantly (p < 0.01) shifted up relative to the control data, a non-parametric Wilcoxon signed-rank test was applied to the data from the treatment and control arrays in a sliding 500-bp window across the genome. A DHS was defined as two or more consecutive positive probes whose central positions are separated by less than 50 bp. Estimated from the negative control probes designed within the Affymetrix microarray, this approach resulted in false positive rates of 0.3% and 0.14% for the 240 U/ml and 480 U/ml DNase I-treated samples from the array readout, respectively. We defined three mutually exclusive DHSs categories; DHSs identified in both samples (875 DHSs), and DHSs only present in one of the two samples treated with either 240 U/ml DNase I (3953 DHSs) or 480 U/ml DNase I (2267 DHSs). The coordinates for all DHSs detected by tiling arrays can be downloaded at .


Genome-scale identification of Caenorhabditis elegans regulatory elements by tiling-array mapping of DNase I hypersensitive sites.

Shi B, Guo X, Wu T, Sheng S, Wang J, Skogerbø G, Zhu X, Chen R - BMC Genomics (2009)

Protocol outline for the genome-scale mapping of C. elegans DHSs by tiling microarray analysis.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651899&req=5

Figure 1: Protocol outline for the genome-scale mapping of C. elegans DHSs by tiling microarray analysis.
Mentions: The protocol for the genome-scale mapping of C. elegans DNase I hypersensitive sites (DHSs) by the tiling microarray is summarized in Figure 1. The Affymetrix C. elegans Tiling 1.0R array contains ~3.2 million 25-mer oligonucleotide probe pairs covering the Watson strand of the entire non-repetitive genome at an average resolution of 25 bp. Synchronized worms in the young adult (YA) stage were treated with floxuridine (FUdR) for more than 8 hours to reduce the background signal from reproduction, without any apparent effect on vitality and longevity [6,7]. Extracted nuclei were digested with different concentrations of DNase I (Figure 2), and samples treated with 240 and 480 U/ml (along with DNA from untreated nuclei) were applied to the tiling assays. The entire procedure was replicated after an interval of about one month, and quantile-normalization was performed on the biological replicates within treatment and control groups [10]. To identify probes that are significantly (p < 0.01) shifted up relative to the control data, a non-parametric Wilcoxon signed-rank test was applied to the data from the treatment and control arrays in a sliding 500-bp window across the genome. A DHS was defined as two or more consecutive positive probes whose central positions are separated by less than 50 bp. Estimated from the negative control probes designed within the Affymetrix microarray, this approach resulted in false positive rates of 0.3% and 0.14% for the 240 U/ml and 480 U/ml DNase I-treated samples from the array readout, respectively. We defined three mutually exclusive DHSs categories; DHSs identified in both samples (875 DHSs), and DHSs only present in one of the two samples treated with either 240 U/ml DNase I (3953 DHSs) or 480 U/ml DNase I (2267 DHSs). The coordinates for all DHSs detected by tiling arrays can be downloaded at .

Bottom Line: Many DHSs appeared to be associated with annotated non-coding RNAs and recently detected transcripts of unknown function.It has been reported that nematode highly conserved non-coding elements were associated with cis-regulatory elements, and we also found that DHSs, particularly distal intergenic DHSs, were significantly enriched in regions that were conserved between the C. elegans and C. briggsae genomes.We describe the first genome-wide analysis of C. elegans DHSs, and show that the distribution of DHSs is strongly associated with functional elements in the genome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bioinformatics Laboratory and National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, PR China. shibaochen@moon.ibp.ac.cn

ABSTRACT

Background: A major goal of post-genomics research is the integrated analysis of genes, regulatory elements and the chromatin architecture on a genome-wide scale. Mapping DNase I hypersensitive sites within the nuclear chromatin is a powerful and well-established method of identifying regulatory element candidates.

Results: Here, we report the first genome-wide analysis of DNase I hypersensitive sites (DHSs) in Caenorhabditis elegans. The data was obtained by hybridizing DNase I-treated and end-captured material from young adult worms to a high-resolution tiling microarray. The data show that C. elegans DHSs were significantly enriched within intergenic regions located 2 kb upstream and downstream of coding genes, and also that a considerable fraction of all DHSs mapped to intergenic positions distant to annotated coding genes. Annotated transcribed loci were generally depleted in DHSs relative to intergenic regions, but DHSs were nonetheless enriched in coding exons and UTRs, whereas introns were significantly depleted in DHSs. Many DHSs appeared to be associated with annotated non-coding RNAs and recently detected transcripts of unknown function. It has been reported that nematode highly conserved non-coding elements were associated with cis-regulatory elements, and we also found that DHSs, particularly distal intergenic DHSs, were significantly enriched in regions that were conserved between the C. elegans and C. briggsae genomes.

Conclusion: We describe the first genome-wide analysis of C. elegans DHSs, and show that the distribution of DHSs is strongly associated with functional elements in the genome.

Show MeSH