In brief, the limbs and tails of mice were cut off, and the skin was carefully peeled. The isolated mouse skin was then incubated in Collagenase type Ⅳ (2mg/ml, Gibco) for 2hrs at 37℃. After incubation, the epiderm was carefully removed from the surface of the skin. The remaining tissues were dissociated and minced into 1mm pieces, which were then resuspended by Dulbecco’s Modified Eagle Medium (DMEM, Hyclone), containing 10% Fetal bovine serum (Hyclone), 2x penicillin/streptomycin (Invitrogen). The isolated skin fibroblasts were collected by flowing through cell strainers.
Growth protocol
Culture in Dulbecco’s Modified Eagle Medium (DMEM, Hyclone), containing 10% Fetal bovine serum (Hyclone), 2x penicillin/streptomycin (Invitrogen). 37℃. The dox-inducible lentiviral system used has been previously described (Maherali et al., 2008). iPSCs were cultured in mESC medium (a 1:1 mixture of Neurolbasal/B27 medium and DMEM/F12 medium supplemented with N2, plus nonessential amino acids, 100 units/ml penicillin, 100 μg/ml streptomycin, 1 mM L-glutamine, 55 μM β-mercaptoethanol, 1 μM PD0325901, and 3 μM CHIR99021) on mitomycin C-treated MEFs. PD0325901 and CHIR99021 could be replaced by LIF (Millipore). The medium was changed every day, and iPSCs were passaged using trypsin (Invitrogen).
Extracted molecule
total RNA
Extraction protocol
Total cellular RNA was extracted with TRIzol reagent (Sigma). Genomic DNA were removed with DNAase(Ambion). Total RNA was purified with RNeasy plus mini kit(Qiagen).
Label
Cy5
Label protocol
Total mRNAs of all samples were labeled with Cy5.
Hybridization protocol
Hybridized to a mouse Oligo Microarray (Phalanx Mouse OneArray® v2, Phalanx Biotech) according to the manufacturer's protocol.
Scan protocol
After hybridization, arrays were scanned using GenePix 4000B scanner (Molecular Devices) and processed using the GenePix Pro 6.0 software (Molecular Devices).
Description
Induced pluripotent stem cells
Data processing
After removing control probes, the data were analyzed according to the manufacturer's protocol (Phalanx Mouse OneArray® v2, Phalanx Biotech). R1_1, R2_2, G3_iPSCs1, G3_iPSCs2, G6_iPSCs1, G6_iPSCs2, MEF_expression_control1, MEF_expression_control2, Dlx3RNAi_iPSCs1, Dlx3RNAi_iPSCs2 were processed together for normalization.
