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High-Throughput Targeted Repeat Element Bisulfite Sequencing (HT-TREBS)

ABSTRACT: In vertebrates, DNA methylation-mediated repression of retrotransposons is essential for the maintenance of genomic integrity. In the current study, we developed a technique termed HT-TREBS (High-Throughput Targeted Repeat Element Bisulfite Sequencing). This technique is designed to measure the DNA methylation levels of individual loci of any repeat families with next-generation sequencing approaches. To test the feasibility of HT-TREBS, we analyzed the DNA methylation levels of the IAPLTR family using a set of 12 different genomic DNA isolated from the brain, liver and kidney of 4 one-week-old littermates of the mouse strain C57BL/6N. This technique has successfully generated the CpG methylation data of 5,233 loci common in all the samples, representing more than 80% of the individual loci of the five targeted subtypes of the IAPLTR family. According to the results, approximately 5% of the IAPLTR loci have less than 80% average CpG methylation levels with no genomic position preference. Further analyses of the IAPLTR loci also revealed the presence of extensive DNA methylation variations between different tissues and individuals. Overall, these data demonstrate the efficiency and robustness of the new technique, HT-TREBS, and also provide new insights regarding the genome-wide DNA methylation patterns of the IAPLTR repeat elements.

ORGANISM(S): Mus musculus

PROVIDER: GSE49222 | GEO | 2014/07/24

SECONDARY ACCESSION(S): PRJNA213307

REPOSITORIES: GEO

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High-Throughput Targeted Repeat Element Bisulfite Sequencing (HT-TREBS)

Project description:In vertebrates, DNA methylation-mediated repression of retrotransposons is essential for the maintenance of genomic integrity. In the current study, we developed a technique termed HT-TREBS (High-Throughput Targeted Repeat Element Bisulfite Sequencing). This technique is designed to measure the DNA methylation levels of individual loci of any repeat families with next-generation sequencing approaches. To test the feasibility of HT-TREBS, we analyzed the DNA methylation levels of the IAPLTR family using a set of 12 different genomic DNA isolated from the brain, liver and kidney of 4 one-week-old littermates of the mouse strain C57BL/6N. This technique has successfully generated the CpG methylation data of 5,233 loci common in all the samples, representing more than 80% of the individual loci of the five targeted subtypes of the IAPLTR family. According to the results, approximately 5% of the IAPLTR loci have less than 80% average CpG methylation levels with no genomic position preference. Further analyses of the IAPLTR loci also revealed the presence of extensive DNA methylation variations between different tissues and individuals. Overall, these data demonstrate the efficiency and robustness of the new technique, HT-TREBS, and also provide new insights regarding the genome-wide DNA methylation patterns of the IAPLTR repeat elements. High-throughput, single-base resolution, singlicate DNA methylation profiles of IAPLTR retrotransposons in the brain, liver , and kidney of four 1-week-old mouse littemates using the developed technique, HT-TREBS.

2014-07-24 | E-GEOD-49222 | biostudies-arrayexpress

DNA methylation variation of human-specific Alu repeats

Project description:DNA methylation is theÂ major repression mechanismÂ for human retrotransposons, such as the Alu family. Here, we haveÂ derivedÂ methylation levels regarding 5238 loci belonging to twoÂ Alu subfamilies, AluYa5 and AluYb8, using High-Throughput Targeted Repeat Element Bisulfite Sequencing (HT-TREBS). The results indicate that ~90% of loci are repressed by high methylation levels. Of the remaining loci, many of these hypomethylated elements are found near gene promoters and show high levels of DNA methylation variation. WeÂ have characterizedÂ this variation in the context of tumorigenesis and inter-individual differences. Comparison of a primary breast tumor and its matched normal tissue revealed early DNA methylation changes in ~1% of AluYb8 elements in response to tumorigenesis. At the same time, AluYa5/Yb8 elements proximal to promoters also showed differences in methylation of up to one order of magnitude even between normalÂ individuals. Overall,Â the current study demonstrates that early loss of methylation occurs during tumorigenesis in a subset of young Alu elements, suggesting their potential clinical relevance. However, techniques such as deep-bisulfite-sequencing of individual loci using HT-TREBS are required to distinguish clinically relevant loci from the background observed forÂ AluYa5/Yb8 elements in general with regard to high levels of inter-individual variation in DNA methylation. HT-TREBS has been used with the Ion Torrent PGM platform to analyze the DNA methylation of 5238 AluYa5/Yb8 elements in a locus-specific manner in human skin-derived fibroblast cells, and a matched normal breast and primary tumor

2016-10-27 | E-GEOD-74420 | biostudies-arrayexpress

DNA methylation variation of human-specific Alu repeats

Project description:DNA methylation is the major repression mechanism for human retrotransposons, such as the Alu family. Here, we have derived methylation levels regarding 5238 loci belonging to two Alu subfamilies, AluYa5 and AluYb8, using High-Throughput Targeted Repeat Element Bisulfite Sequencing (HT-TREBS). The results indicate that ~90% of loci are repressed by high methylation levels. Of the remaining loci, many of these hypomethylated elements are found near gene promoters and show high levels of DNA methylation variation. We have characterized this variation in the context of tumorigenesis and inter-individual differences. Comparison of a primary breast tumor and its matched normal tissue revealed early DNA methylation changes in ~1% of AluYb8 elements in response to tumorigenesis. At the same time, AluYa5/Yb8 elements proximal to promoters also showed differences in methylation of up to one order of magnitude even between normal individuals. Overall, the current study demonstrates that early loss of methylation occurs during tumorigenesis in a subset of young Alu elements, suggesting their potential clinical relevance. However, techniques such as deep-bisulfite-sequencing of individual loci using HT-TREBS are required to distinguish clinically relevant loci from the background observed for AluYa5/Yb8 elements in general with regard to high levels of inter-individual variation in DNA methylation.

2016-10-27 | GSE74420 | GEO

DNA methylation in Breast cancer

Project description:This experiment was performed with an aim to establish an early diagnostic marker for breast cancer based on the DNA methylation patterns. CpG Island were identified and amplified from the breast cancer related genomic loci. These selected loci were amplified using PCR from different human breast tissue samples. The samples included Breast cancer tissues and macroscopically normal tissues (3-5cm away from tumor) from breast cancer patients, benign breast tissues and healthy breast tissues from cancer free patients. All the genomic loci amplified from one particular tissue sample were pooled, biotin labeled and hybridized to a self designed and synthesised oligonucleotide microarray. The obtained signals were statistically analysed and the genomic loci which could differentiate the different kinds of breast tissues used based on their methylation levels are being reported.

2011-12-30 | E-TABM-1171 | biostudies-arrayexpress

Detection of aberrant DNA methylation in colorectal carcinoma samples compared to normal human colon

Project description:To globally define methylation-âproneâ and -âprotectedâ CpG islands in colorectal carcinoma we analyzed the methylation status of 23,000 CpG islands of the human genome in ten coleorectal carcinoma samples as well as normal colon using our previously described methyl-CpG immunoprecipitation (MCIp) technique (Gebhard et al. 2006; Schilling and Rehli 2007). This method enriches for highly CpG methylated DNA that can be directly applied to fluorescent labeling and oligonucleotide microarray hybridization without an additional amplification step. Keywords: MCIp-on-Chip; comparative genomic hybridization CpG-methylated genomic DNA was enriched using methyl-CpG immunoprecipitation (MCIp). On each microarray, the enriched material from colorectal carcinoma samples was compared to the enriched material from normal colon to identify aberrantly methylated regions.

2010-06-05 | E-GEOD-17512 | biostudies-arrayexpress

CpG context and content in evolutionary signatures of brain DNA methylation

Project description:We assayed CpG methylation in cerebral cortex of neurologically and psychiatrically normal human postmortem specimens, as well as mouse forebrain specimens. Cross-species human-mouse DNA methylation conservation analysis shows that DNA methylation is not correlated with sequence conservation. Instead, greater DNA methylation conservation is correlated with increasing CpG density. We identified key genomic features that can be targeted for identification of epigenetic loci that may be developmentally and evolutionarily conserved and wherein aberrations in DNA methylation patterns can confer risk for disease. Characterization of evolutionary signatures of DNA methylation in the brain

2011-10-04 | E-GEOD-32647 | biostudies-arrayexpress

CpG context and content in evolutionary signatures of brain DNA methylation

2011-10-05 | GSE32647 | GEO

Detection of aberrant DNA methylation in acute leukemia samples compared to normal human monocytes

Project description:To globally define methylation-âproneâ and -âprotectedâ CpG islands in leukemia, we analyzed the methylation status of 23,000 CpG islands of the human genome in eight acute leukemia samples as well as normal blood monocytes using our previously described methyl-CpG immunoprecipitation (MCIp) technique (Gebhard et al. 2006; Schilling and Rehli 2007). This method enriches for highly CpG methylated DNA that can be directly applied to fluorescent labeling and oligonucleotide microarray hybridization without an additional amplification step. Keywords: MCIp-on-Chip; comparative genomic hybridization CpG-methylated genomic DNA was enriched using methyl-CpG immunoprecipitation (MCIp). On each microarray, the enriched material from leukemia samples was compared to the enriched material from normal blood monocytes to identify aberrantly methylated regions. Three biological replicates were analysed for each cell line.

2010-06-05 | E-GEOD-17510 | biostudies-arrayexpress

Detection of aberrant DNA methylation in AML cell lines compared to normal human monocytes

Project description:To globally define methylation-âproneâ and -âprotectedâ CpG islands in leukemia, we analyzed the methylation status of 23,000 CpG islands of the human genome in two acute leukemia cell lines as well as normal blood monocytes using our previously described methyl-CpG immunoprecipitation (MCIp) technique (Gebhard et al. 2006; Schilling and Rehli 2007). This method enriches for highly CpG methylated DNA that can be directly applied to fluorescent labeling and oligonucleotide microarray hybridization without an additional amplification step. Keywords: MCIp-on-Chip; comparative genomic hybridization CpG-methylated genomic DNA was enriched using methyl-CpG immunoprecipitation (MCIp). On each microarray, the enriched material from leukemia cell lines was compared to the enriched material from normal blood monocytes to identify aberrantly methylated regions. Three biological replicates were analysed for each cell line.

2010-06-05 | E-GEOD-17455 | biostudies-arrayexpress

Detection of aberrant DNA methylation in colorectal carcinoma samples compared to normal human colon

Project description:To globally define methylation-’prone’ and -’protected’ CpG islands in colorectal carcinoma we analyzed the methylation status of 23,000 CpG islands of the human genome in ten coleorectal carcinoma samples as well as normal colon using our previously described methyl-CpG immunoprecipitation (MCIp) technique (Gebhard et al. 2006; Schilling and Rehli 2007). This method enriches for highly CpG methylated DNA that can be directly applied to fluorescent labeling and oligonucleotide microarray hybridization without an additional amplification step. Keywords: MCIp-on-Chip; comparative genomic hybridization

2010-03-31 | GSE17512 | GEO

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