GSM2844763: iPD mouse4; Mus musculus; RNA-Seq
Generation of Arid1aflox/flox mice on the C57BL/6 background was previously described (Gao et al., 2008). Cre-mediated deletion of Arid1a removed exon 8 and introduced a frameshift mutation and a premature stop codon (p.Gly809Hisfs*6; wild type ARID1A protein has 2283 amino acids). Ptenflox/flox mice on the BALB/c background (Strain C;129S4-Ptentm1Hwu/J) were obtained from the Jackson Laboratory (Groszer et al., 2001). Cre-mediated deletion of Pten removed exon 5 and introduced a frameshift mutation (p.Val85Glyfs*14). Arid1aflox/flox;Ptenflox/flox mice were generated by intercrossing these transgenic strains. In order to express Cre recombinase specifically in the mouse uterine epithelium for all subsequent genetic alterations described, we used Pax8-Cre mice which were generated by crossing mice expressing the reverse tetracycline-controlled transactivator (rtTA) under the control of the Pax8 promoter (Pax8-rtTA) with mice expressing Cre recombinase in a tetracycline-dependent manner (TetO-Cre) (Perets et al., 2013).  To generate mouse models with Arid1a and Pten individual or combined knockout in uterine epithelium, we crossed Pax8-Cre mice with Arid1aflox/flox mice, Ptenflox/flox mice, and Arid1aflox/flox;Ptenflox/flox mice. Figure S4A summarizes the generation of these genetically engineered mice in which Arid1a and Pten can be conditionally knocked out. All experimental mice were maintained on a mixed genetic background (C57BL/6, BALB/c, and S129). The resultant genotype of each model was confirmed by genomic DNA PCR, using primer sequences described previously (Guan et al., 2014). The wild-type, floxed, and recombined Arid1a generate 669, 845, and 298 bp PCR products, respectively. The predicted PCR products for wild-type, flox, and recombined Pten are 1018, 1100, and 400 bp, respectively. Knockout was initiated by treating mice with doxycycline either through oral gavage (2 mg/mouse/day). All of the animal procedures were approved by the Johns Hopkins University Animal Care Committee. Uteri from 4 iPAD and 4 iPD mice were harvested two weeks after gene knockout (by feeding doxycycline). Hematoxylin-and-eosin stained tissue sections were prepared, and the diagnosis of endometrioid carcinoma from all iPAD mice and EIN from all iPD mice was confirmed by histopathology. Total RNA was isolated from mouse uterine tissues using the Qiagen RNeasy Plus Mini Kit. RNA quality was assessed using the Agilent 2100 Bioanalyzer RNA Nano Chip. RIN values ranged from 5.8 to 7.8. RNA-sequencing was performed by GeneWiz, Inc. on the Illumina HiSeq2500 platform, in a 2x100 bp paired-end high output V4 chemistry configuration.
GEO Accession
GSM2844763