Genomic DNA from MEF cells

ABSTRACT: Genome wide DNA methylation profiling of WT and Pml KO mEF cells treated W/O Doxorubicin for 24 hours. TET2 plays critical roles in the regulation of cell identity and inhibition of tumorigenesis through modulating DNA methylation. However, little is known about how TET2 is targeted to specific DNA-binding sites. It was found by us that TET2 is significantly recruited to specific sites by PML. Moreover, TET2 and PML synergistically inhibited cell proliferation. Further study indicated that multi chemotherapies, such as doxorubicin, were able to promote PML-NB, increase 5hmC and result in specific DNA demethylation. Due to the important role of inactivation of tumor suppressors by DNA methylation in cancer, the activity of most therapies depends to an extent on activation of tumor suppressors by demethylation. Consequently, it is not surprising that methylation of tumor suppressors has been reported to diminish in responder patients after end of chemotherapy. Little is known about mechanism of therapy-promoted DNA demethylation, although this therapy-promoted DNA demethylation potentially plays general roles in therapy response, resistance and recurrence. It was also found that knock out of Pml significantly diminished chemotherapy-promoted DNA demethylation. To identify PML-TET2 pathway-regulated genes, WT and Pml knock out MEF cells were treated W/O Doxorubicin. The genomic DNA was extracted and purified from samples using Qiagen DNeasy Kit according to manufacturerâ??s instruction. The resultant Genome DNA was cut by Restriction Enzyme. After DNA-end repair and 3â??-dA overhang, the 40-220bp fragments were selected and treated with ZYMO EZ DNA Methylation-Gold kit according to manufacturerâ??s instruction. Then the bisulphite converted DNA from the 4 samples (WT, WT Dox, PML KO and Pml KO Dox) were sequenced using standard Illumina protocol. Standard recommended Illumina scanner setting was carried out in study. WT and Pml knock out MEF cells were treated with Doxorubicin. The genomic DNA was extracted and purified from samples using Qiagen DNeasy Kit according to manufacturerâ??s instruction. The resultant Genome DNA was cut by Restriction Enzyme. After DNA-end repair and 3â??-dA overhang, the 40-220bp fragments were selected and treated with ZYMO EZ DNA Methylation-Gold kit according to manufacturerâ??s instruction. Then the bisulphite converted DNA from the 4 samples (WT, WT Dox, PML KO and Pml KO Dox) were sequenced using standard Illumina protocol. Standard recommended Illumina scanner setting was carried out in study.

ORGANISM(S): Mus musculus

SUBMITTER: Qingkai Yang 

PROVIDER: E-GEOD-75408 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress