|
Status |
Public on Dec 01, 2012 |
Title |
RRBS HL60 rep1 |
Sample type |
SRA |
|
|
Source name |
HL60 Cell lines
|
Organism |
Homo sapiens |
Characteristics |
cell line: HL60 digestion: MspI treatment: bisulfite
|
Treatment protocol |
Genomic DNA was isolated from the myeloid cell lines HL-60. DNA was digested with MspI (NEB), a methylation-insensitive enzyme that cuts C'CGG. Digested DNA was size selected on a 4% NuSieve 3:1 Agarose gel (Lonza). For each sample, two slices containing DNA fragments of 40-120 bp and 120-220 bp, respectively, were excised from the unstained preparative portion of the gel. These two size fractions were kept apart throughout the procedure including the final sequencing. Pre-annealed Illumina adaptors containing 5'-methyl-cytosine instead of cytosine were ligated to size-selected MspI fragments. Adapter-ligated fragments were bisulfite-treated using the EZ DNA Methylation kit (Zymo Research, Orange, CA). The products were PCR amplified, size selected, and sequenced on the Illumina GA IIxat a reading length of 36 bp.
|
Growth protocol |
HL-60 were grown in DMEM supplemented with 10%FBS, at 37 °C in a humidified atmosphere with 5% CO2
|
Extracted molecule |
genomic DNA |
Extraction protocol |
DNA was digested with MspI (NEB), a methylation-insensitive enzyme that cuts C'CGG. Digested DNA was size selected on a 4% NuSieve 3:1 Agarose gel (Lonza). For each sample, two slices containing DNA fragments of 40-120 bp and 120-220 bp, respectively, were excised from the unstained preparative portion of the gel. These two size fractions were kept apart throughout the procedure including the final sequencing. Pre-annealed Illumina adaptors containing 5'-methyl-cytosine instead of cytosine were ligated to size-selected MspI fragments. Adapter-ligated fragments were bisulfite-treated using the EZ DNA Methylation kit (Zymo Research, Orange, CA). The products were PCR amplified, size selected before subjecting to sequencing.
|
|
|
Library strategy |
Bisulfite-Seq |
Library source |
genomic |
Library selection |
Reduced Representation |
Instrument model |
Illumina Genome Analyzer IIx |
|
|
Description |
Reduced Representation Bisulfite Sequencing of HL60
|
Data processing |
Alignment: Sequencing reads were mapped to the reference genome hg19 using RRBSmap allowing 2 mismatches.
|
|
|
Submission date |
Sep 15, 2011 |
Last update date |
May 15, 2019 |
Contact name |
Touati Benoukraf |
E-mail(s) |
tbenoukraf@mun.ca
|
Phone |
+1 (709) 864-6671
|
Organization name |
Memorial University of Newfoundland
|
Department |
Faculty of Medicine, Division of BioMedical Sciences
|
Street address |
Craig L. Dobbin Genetics Research Centre,
|
City |
St. John's |
State/province |
NL |
ZIP/Postal code |
A1B 3V6 |
Country |
Canada |
|
|
Platform ID |
GPL10999 |
Series (2) |
GSE32168 |
RRBS (Reduced Representation Bisulfite Sequencing) of HL60 |
GSE32260 |
Relationship between DNMT1-RNA interactions, DNA methylation and gene expression |
|
Relations |
SRA |
SRX103284 |
BioSample |
SAMN00744409 |