|
| Status |
Public on Oct 05, 2023 |
| Title |
Extraembryonic mesoderm fetal biopsy [PL2409_EM3584 methylation] |
| Sample type |
genomic |
| |
|
| Source name |
Extraembryonic mesoderm
|
| Organism |
Homo sapiens |
| Characteristics |
gender: Female
tissue: Extraembryonic mesoderm fetal biopsy
|
| Treatment protocol |
The two tissues were mechanically separated and the yield of chorionic cytotrophoblast cells was increased by maceration of chorionic villi under an Axiovert 200 inverted microscope (Carl Zeiss, Germany) and treatment with 70% acetic acid for 3-5 min followed by three washes of the obtained cell suspension with PBS according to a modified protocol (Simoni, G. et al. Efficient direct chromosome analyses and enzyme determinations from chorionic villi samples in the first trimester of pregnancy. Human Genetics 63, 349-357 (1983)).
|
| Growth protocol |
The tissues were chopped with scissors to small fragments, and long-term cultures were set up in 25 cm2 flasks with 5 mL of DMEM/F12 (1:1) medium (Gibco) supplemented with 20% fetal bovine serum (HyClone), 1% MEM NEAA solution (Gibco), 1% Pen-Strep (Gibco). Tissues were incubated at 37°C with once-weekly medium renewal.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted using a standard phenol-chloroform extraction method that allows for the isolation of up to 900 ng DNA from the tissues. Likewise, genomic DNA was extracted from peripheral blood of parents using the standard phenol-chloroform method.
|
| Label |
cy3, cy5
|
| Label protocol |
Standard Illumina Protocol
|
| |
|
| Hybridization protocol |
From each sample 300 ng of DNA was bisulfite-converted using the EZ DNA Methylation Kit (Zymo Research, CA, USA) according to the manufacturer’s instructions
|
| Scan protocol |
Illumina Infinium™ MethylationEPIC v1.0 BeadChip (Illumina, CA, USA).
|
| Description |
PL2409_EM3584
|
| Data processing |
Data preprocessing was carried out using the RnBeads R package. Briefly, the data were normalized with subset-quantile within array normalization (SWAN) and poor-quality sites/samples were removed based on the greedycut algorithm (detection p-value threshold: 0.05). Further sites were removed: (i) sites on the sex chromosomes, (ii) sites in close proximity to SNPs, (iii) sites with missing values in more than 10% of samples, (iv) sites not in a CpG context. Additional sample quality control, namely sex prediction and SNP probe analysis, was carried out using the sEst42 package and RnBeads respectively.
|
| |
|
| Submission date |
Mar 24, 2023 |
| Last update date |
Oct 05, 2023 |
| Contact name |
Rick Essers |
| E-mail(s) |
rick.essers@mumc.nl
|
| Organization name |
Maastricht University Medical Centre (MUMC+)
|
| Department |
Clinical Genetics
|
| Street address |
P. Debyelaan 25
|
| City |
Maastricht |
| State/province |
Limburg |
| ZIP/Postal code |
6229HX |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL21145 |
| Series (2) |
| GSE228149 |
Prevalence of chromosomal alterations in first-trimester spontaneous pregnancy loss |
| GSE228151 |
Prevalence of chromosomal alterations in first-trimester spontaneous pregnancy loss |
|
