Purification of CpG islands using a methylated DNA binding column

Cross, Sally H.; Charlton, Jillian; Nan, Xinsheng; Bird, Adrian

doi:10.1038/ng0394-236

Cited by 418 publications

(307 citation statements)

References 40 publications

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“…Our sequence analysis of HSulf-1 promoter revealed putative CpG-rich sequences in Exon 1A (Lai et al, 2003). This fragment (Z58846) was also identified as a putative CpG-rich sequence in previous study (Cross et al, 1994). In this study, we report on the methylation status of this putative CpG island in ovarian cell lines and primary ovarian tumors by direct sequencing of the bisulfitemodified DNA.…”

Section: Introductionsupporting

confidence: 64%

“…Methylation-associated silencing of HSulf-1 in ovarian cell lines and primary ovarian tumors Primers encompassing 12 CpG sites within the CpG-rich region in Exon 1A of HSulf-1 (Cross et al, 1994;Lai et al, 2003) were designed to amplify and sequence the bisulfite-modified DNA as described previously (Herman et al, 1996;Shridhar et al, 2001). Overall methylation was determined for a subset of cell lines and primary ovarian tumors (fresh frozen) with or without HSulf-1 expression.…”

Section: Resultsmentioning

confidence: 99%

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Epigenetic silencing of HSulf-1 in ovarian cancer:implications in chemoresistance

et al. 2007

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To investigate the mechanism by which HSulf-1 expression is downregulated in ovarian cancer, DNA methylation and histone acetylation of HSulf-1 was analysed in ovarian cancer cell lines and primary tumors. Treatment of OV207 and SKOV3 by 5-aza-2 0 -deoxycytidine resulted in increased transcription of HSulf-1. Sequence analysis of bisulfite-modified genomic DNA from ovarian cell lines and primary tumors without HSulf-1 expression revealed an increase in the frequency of methylation of 12 CpG sites in exon 1A. Chromatin immunoprecipitation assays showed an increase in histone H3 methylation in cell lines without HSulf-1 expression. To assess the significance of HSulf-1 downregulation in ovarian cancer, OV167 and OV202 cells were transfected with HSulf-1 siRNA. Downregulation of HSulf-1 expression in OV167 and OV202 cells lead to an attenuation of cisplatin-induced cytotoxicity. Moreover, patients with ovarian tumors expressing higher levels of HSulf-1 showed a 90% response rate (27/30) to chemotherapy compared to a response rate of 63% (19/30) in those with weak or moderate levels (P ¼ 0.0146, v 2 test). Collectively, these data indicate that HSulf-1 is epigenetically silenced in ovarian cancer and that epigenetic therapy targeting HSulf-1 might sensitize ovarian tumors to conventional first-line therapies.

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Section: Introductionsupporting

confidence: 64%

Section: Resultsmentioning

confidence: 99%

Epigenetic silencing of HSulf-1 in ovarian cancer:implications in chemoresistance

et al. 2007

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“…A panel of 12 288 CpG island clones was created from the CGI genomic library from Cross et al, 1994, as part of the Human Genome Mapping Project Resource Center (Heisler et al, 2005). Inserts from this CGI library were polymerase chain reaction (PCR)-amplified, purified by ethanol precipitation and then arrayed onto glass microscope slides by a specialised robot designed and created at Albert Einstein College of Medicine (Cheung et al, 1999;Adrien et al, 2006).…”

Section: Cpg Island Arraysmentioning

confidence: 99%

Identification of DNA hypermethylation of SOX9 in association with bladder cancer progression using CpG microarrays

et al. 2007

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CpG island arrays represent a high-throughput epigenomic discovery platform to identify global disease-specific promoter hypermethylation candidates along bladder cancer progression. DNA obtained from 10 pairs of invasive bladder tumours were profiled vs their respective normal urothelium using differential methylation hybridisation on custom-made CpG arrays (n ¼ 12 288 clones). Promoter hypermethylation of 84 clones was simultaneously shown in at least 70% of the tumours. SOX9 was selected for further validation by bisulphite genomic sequencing and methylation-specific polymerase chain reaction in bladder cancer cells (n ¼ 11) and primary bladder tumours (n ¼ 101). Hypermethylation was observed in bladder cancer cells and associated with lack of gene expression, being restored in vitro by a demethylating agent. In primary bladder tumours, SOX9 hypermethylation was present in 56.4% of the cases. Moreover, SOX9 hypermethylation was significantly associated with tumour grade and overall survival. Thus, this high-throughput epigenomic strategy has served to identify novel hypermethylated candidates in bladder cancer. In vitro analyses supported the role of methylation in silencing SOX9 gene. The association of SOX9 hypermethylation with tumour progression and clinical outcome suggests its relevant clinical implications at stratifying patients affected with bladder cancer.

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“…This requires efficient strategies for the detection of 5mC. Different concepts for discriminating between cytosine (C) and 5mC have been described which rely on endonuclease digestion,8 affinity enrichment,9 nanopore sequencing,10 different chemical behavior concerning redox reactivity,11 or selective deamination of C using sodium bisulfite 12…”

mentioning

confidence: 99%

Modified Nucleotides for Discrimination between Cytosine and the Epigenetic Marker 5‐Methylcytosine

2016

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Abstract5‐Methyl‐2′‐deoxycytosine, the most common epigenetic marker of DNA in eukaryotic cells, plays a key role in gene regulation and affects various cellular processes such as development and carcinogenesis. Therefore, the detection of 5mC can serve as an important biomarker for diagnostics. Here we describe that modified dGTP analogues as well as modified primers are able to sense the presence or absence of a single methylation of C, even though this modification does not interfere directly with Watson–Crick nucleobase pairing. By screening several modified nucleotide scaffolds, O6‐modified 2′‐deoxyguanosine analogues were identified as discriminating between C and 5mC. These modified nucleotides might find application in site‐specific 5mC detection, for example, through real‐time PCR approaches.

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Purification of CpG islands using a methylated DNA binding column

Cited by 418 publications

References 40 publications

Epigenetic silencing of HSulf-1 in ovarian cancer:implications in chemoresistance

Epigenetic silencing of HSulf-1 in ovarian cancer:implications in chemoresistance

Identification of DNA hypermethylation of SOX9 in association with bladder cancer progression using CpG microarrays

Modified Nucleotides for Discrimination between Cytosine and the Epigenetic Marker 5‐Methylcytosine

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