|Public on May 06, 2022
|mouse embryonic fibroblasts
|cell type: mouse embryonic fibroblasts
treatment: mouse embryonic fibroblasts after 10 days of Repro
treatment time: 10 days
|MEF are infected by OSKM retroviruses to induce Repro or by K-Ras G12V, shp53 lentiviruses combined with cMyc retroviral infection to induce Transfo.
|MEF were generated from E13.5 Bcl11b-tdTomato mice embryos and cultivated in DMEM medium supplemented with 10% foetal bovine serum (FBS), 100U/ml penicillin / streptomycin (PS), 1mM sodium pyruvate, 2mM L-glutamine, 0.1mM Non Essential Amino Acids (NEAA) and 0.1mM b-mercaptoethanol. iPS cells were grown in DMEM containing 15% KnockOut™ Serum Replacement, 1,000 U/ml leukemia inhibitory factor, 100U/ml PS, 1mM sodium pyruvate, 2mM L-glutamine, 0.1mM NEAA and 0.1mM b-mercaptoethanol. Transformed cells were cultivated in the same medium than MEF.
|Cell suspension was mixed with master mix and loaded with Single Cell 3′ Gel Beads and Partitioning Oil into a Single Cell G Chip. RNA transcripts from single cells were uniquely barcoded and reverse-transcribed within droplets. cDNA molecules were pooled and the cDNA pool then go through an end repair process, the addition of a single ‘A’ base, and then ligation of the adapters. The products are then purified and enriched with PCR to create the final cDNA library.
10x Genomics Chromium 3' v3.1 Single Cell Gene Expression Kit protocol was followed according to manufacturer instructions.
|Illumina NovaSeq 6000
|Bcl2 files were transormed to fastq files with cellranger mkfastq v 5.0.1.
Alignment, filtering and quantification were done with cellranger count v 5.0.1.
Supplementary_files_format_and_content: Outputs from cellranger count: barcodes, features and matrix files.
|Aug 04, 2021
|Last update date
|May 06, 2022
|28 rue Laennec
|The comparative roadmaps of reprogramming and transformation unveiled that cellular plasticity is broadly controlled by Bcl11b and Atoh8
|Cellular identity loss occurring during both iPS cells generation and malignant transformation can be followed using the somatic markers Thy1 and Bcl11b [scRNA-seq]