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| Status |
Public on Sep 04, 2023 |
| Title |
Quartet_methy_ILM_850K_SNT_D6_CGZ302_6993610 |
| Sample type |
genomic |
| |
|
| Source name |
genomic DNA from B lymphocyte
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: D6
Sex: Female
disease state: normal
cell type: immortalized B-lymphoblastoid cell lines
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| Treatment protocol |
To obtain the first batch of DNA reference materials (Lot NO 20160806), 2 x109 cells were harvested simultaneously for each cell line. Specifically, the cellsgrew in suspension and were centrifuged at 300 g for 5 mins to obtain celpellets. The cell pellets were then washed twice with cold PBS. The DNA reference materials were purified using the DNA with Blood &Cell Culture DNA Maxi Kit (Qiagen, Germany) according to the manufacturer'sinstructions, divided into 1000 aliquots for each of the Quartet members, andthen labeled as Quartet DNA D5 20160806, Quartet DNA D6 2016080641Quartet DNA F7 20160806, and Quartet DNA M8 20160806. A single viacontains approximate 10 ug of genomic DNA (220 ng/uL, 50 uL) in TE buffer(10 mM TRIS, pH 8.0, 1 mM EDTA, pH 8.0).The DNA integrity and long-term stability were evaluated by Agilent 2200TapeStation system (Agilent Technologies, USA). Concentrations weredetermined by NanoDrop ND-2000 spectrophotometer (Thermo FisherScientific,USA).
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| Growth protocol |
Immortalized lymphoblastoid cell lines were established by Epstein-Barr virus (EBV). Peripheral blood mononuclear cells were isolated using a lymphocyte separation solution (Ficoll). Naïve B cells were sorted by EasySep Human naïve B Cell Enrichment Kit (STEMCELL, Catalog#19254), and infected by Epstein-Barr virus (EBV) by centrifugation at 2000 rpm for 1 hour. After incubation, the successfully infected and immortalized cells were propagated in culture medium. The Quartet LCLs were cultured in RPMI 1640 with 2 mM L-glutamine, 10% heat-inactivated FBS (fetal bovine serum), and 1% PS (penicillin/streptomycin) at 37 °C with 5% CO2. The cells were passaged every 72 hours at a 1:4 split ratio.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
genomic DNA was extracted and purified from LBC samples using Qiagen DNeasy Kit according to standard instructions
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| Label |
Cy5 and Cy3
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| Label protocol |
Standard Illumina Protocol
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| |
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| Hybridization protocol |
Bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC BeadChip using standard Illumina protocol
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| Scan protocol |
Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
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| Data processing |
Raw idat files were processed using R package ChAMP v2.20.1. Next, samples with a proportion of failed probes (probe detection p-value > 0.01) above 0.1 were discarded. Finally, the Meth and Unmeth signals were used to calculate M values and β values. In this process, the offset was set to 100 and the beta threshold to 0.001.
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| |
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| Submission date |
Aug 30, 2023 |
| Last update date |
Sep 04, 2023 |
| Contact name |
Leming Shi |
| E-mail(s) |
[email protected]
|
| Phone |
+86-18616827008
|
| Organization name |
Fudan University
|
| Department |
School of Life Sciences
|
| Lab |
Center for Pharmacogenomics
|
| Street address |
2005 Songhu Road
|
| City |
Shanghai |
| ZIP/Postal code |
200438 |
| Country |
China |
| |
|
| Platform ID |
GPL19718 |
| Series (1) |
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