|
| Status |
Public on Nov 23, 2009 |
| Title |
Normal bone marrow, 629 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
bone marrow, normal
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: bone marrow
gender: female
genotype: normal
|
| Treatment protocol |
Only applicable for the cell lines: treated with 100nM of zebularine for 10 days.
|
| Growth protocol |
All cell lines were maintained as suspension cultures in RPMI 1640 with L-Alanyl-L-Glutamine (Invitrogen) supplemented with 10% FCS (Integro), 100 IU/ml penicillin, 100 μg/ml streptomycin, and 0.125 μg/ml fungizone (Invitrogen) at 37°C in humidified air containing 5% CO2.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Total DNA extracted using Trizol following manufacturer's instructions.
|
| Label |
Cy5
|
| Label protocol |
Using the BioPrime Array-CGH Genomic Labeling kit (Invitrogen, Carlsbad, USA), amino-allyl dUTPs were incorporated into 500ng of the methylated amplicons (generated by DMH) , allowing the amplicons to be labeled with the fluorescent dyes Cy5 (patient samples) and Cy3 (common reference samples).
|
| |
|
| Channel 2 |
| Source name |
peripheral blood, healthy
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: peripheral blood
sample description: mix from 5 healthy males and 5 healthy females
sample type: reference
|
| Treatment protocol |
Only applicable for the cell lines: treated with 100nM of zebularine for 10 days.
|
| Growth protocol |
All cell lines were maintained as suspension cultures in RPMI 1640 with L-Alanyl-L-Glutamine (Invitrogen) supplemented with 10% FCS (Integro), 100 IU/ml penicillin, 100 μg/ml streptomycin, and 0.125 μg/ml fungizone (Invitrogen) at 37°C in humidified air containing 5% CO2.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Total DNA extracted using Trizol following manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
Using the BioPrime Array-CGH Genomic Labeling kit (Invitrogen, Carlsbad, USA), amino-allyl dUTPs were incorporated into 500ng of the methylated amplicons (generated by DMH) , allowing the amplicons to be labeled with the fluorescent dyes Cy5 (patient samples) and Cy3 (common reference samples).
|
| |
|
| |
| Hybridization protocol |
Oligoarray control targets and hybridization buffer (the Oligo aCGH/ChIP-on-Chip Hybridization Kit) were added, and samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequentially according to the Agilent protocol.
|
| Scan protocol |
Scanned on an Agilent G2565AA scanner.
|
| Description |
Please refer to Differential Methylation Hybridization (DMH) protocol by Dr. Tim H. Huang and coworkers. (PMID 18987809: Yan PS, Potter D, Deatherage DE, Huang TH, Lin S., Differential methylation hybridization: profiling DNA methylation with a high-density CpG island microarray.Methods Mol Biol. 2009;507:89-106.)
|
| Data processing |
Images were quantified using Agilent Feature Extraction Software (version A.8.5.1.1).
Agilent Feature Extraction Software (v 8.5.1.1) was used for data extraction. The R and Bioconductor statistical environment (version 2.7) were used for quality control and LOESS normalization (limma package). No background correction was performed.
|
| |
|
| Submission date |
Oct 02, 2009 |
| Last update date |
Nov 23, 2009 |
| Contact name |
Dominique Stumpel |
| Organization name |
Erasmus MC _ Sophia Children's Hospital
|
| Department |
Pediatric Oncology/ Hematology
|
| Lab |
Pediatric Oncology/ Hematology
|
| Street address |
Dr. Molewaterplein 50-room Ee15-14
|
| City |
Rotterdam |
| ZIP/Postal code |
3061KS |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL4126 |
| Series (1) |
|
