|
| Status |
Public on Sep 01, 2012 |
| Title |
M1+IL6 H3K27me3 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
M1+IL6 H3K27me3
|
| Organism |
Mus musculus |
| Characteristics |
cell line: M1+IL-6
c-myb expression: No
antibody: H3K27me3
|
| Treatment protocol |
For cell differentiation, M1 cells were seeded at a density of 1-2 x 105 cells / ml in medium containing interleukin-6 (IL-6) for 4-5 days.
|
| Growth protocol |
M1cells were seeded in RPMI 1640 medium with 10% (v/v) heat-inactivated horse serum and treated with interleukin-6 (IL-6) for5 days
M1cells were seeded in RPMI 1640 medium with 10% (v/v) heat-inactivated horse serum and treated with interleukin-6 (IL-6) for5 days
The murine myeloid cell line M1 was maintained in RPMI 1640 medium with 10% (v/v) heat-inactivated horse serum (Invitrogen). All tumor cell lines were cultured in Dulbecco modified Eagle medium with 10% (v/v) fetal bovine serum.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin immunoprecipitation assays was conducted as previously described (Paul, Bies et al. 2010). Cells were fixed in 0.8% formaldehyde for 6 minutes at room temperature. After lysis, samples were sonicated to a size range of 200 to 1000 bp. Chromatin (150-200 µg) was immunoprecipitated with antibodies for H3K4me3 (Abcam, ab8580), H3K4me1 (Abcam, ab8895), H3K9/14Ac (Upstate, 06-599), CTCF (Abcam, ab70303), H3K9me3 (Abcam, ab8898), H3K27me3 (Upstate,17-622),or rabbit IgG (Sigma-Aldrich, 15006). A 10% aliquot was removed as an input fraction. ChIP DNA and input DNA were amplified using WGA2 kit (Sigma-Aldrich).
|
| Label |
Cy5
|
| Label protocol |
A total of 2.5 µg of amplified DNA was labeled with Cy3 (input) or Cy5 (IP) dUTP (PerkinElmer Life and Analytical Sciences) using the CGH labeling kit (Invitrogen).
|
| |
|
| Channel 2 |
| Source name |
M1+IL6 Input
|
| Organism |
Mus musculus |
| Characteristics |
cell line: M1+IL-6
c-myb expression: No
|
| Treatment protocol |
For cell differentiation, M1 cells were seeded at a density of 1-2 x 105 cells / ml in medium containing interleukin-6 (IL-6) for 4-5 days.
|
| Growth protocol |
M1cells were seeded in RPMI 1640 medium with 10% (v/v) heat-inactivated horse serum and treated with interleukin-6 (IL-6) for5 days
M1cells were seeded in RPMI 1640 medium with 10% (v/v) heat-inactivated horse serum and treated with interleukin-6 (IL-6) for5 days
The murine myeloid cell line M1 was maintained in RPMI 1640 medium with 10% (v/v) heat-inactivated horse serum (Invitrogen). All tumor cell lines were cultured in Dulbecco modified Eagle medium with 10% (v/v) fetal bovine serum.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin immunoprecipitation assays was conducted as previously described (Paul, Bies et al. 2010). Cells were fixed in 0.8% formaldehyde for 6 minutes at room temperature. After lysis, samples were sonicated to a size range of 200 to 1000 bp. Chromatin (150-200 µg) was immunoprecipitated with antibodies for H3K4me3 (Abcam, ab8580), H3K4me1 (Abcam, ab8895), H3K9/14Ac (Upstate, 06-599), CTCF (Abcam, ab70303), H3K9me3 (Abcam, ab8898), H3K27me3 (Upstate,17-622),or rabbit IgG (Sigma-Aldrich, 15006). A 10% aliquot was removed as an input fraction. ChIP DNA and input DNA were amplified using WGA2 kit (Sigma-Aldrich).
|
| Label |
Cy3
|
| Label protocol |
A total of 2.5 µg of amplified DNA was labeled with Cy3 (input) or Cy5 (IP) dUTP (PerkinElmer Life and Analytical Sciences) using the CGH labeling kit (Invitrogen).
|
| |
|
| |
| Hybridization protocol |
A total of 3 µg of labeled ChIP and input DNA was cohybridized to the chip for 40 hours at 65°C
|
| Scan protocol |
scanned on an Agilent Scanner with Agilent Scan Control 7.0 software
|
| Description |
M1 cells treated with IL-6 for 5 days
|
| Data processing |
Data was extracted with Feature Extraction 9.1 software and analyzed using ChIP Analytics 1.3 software (Agilent).
|
| |
|
| Submission date |
Dec 29, 2011 |
| Last update date |
Sep 01, 2012 |
| Contact name |
Junfang Zhang |
| E-mail(s) |
zhangj3@mail.nih.gov
|
| Phone |
301-402-9232
|
| Fax |
301-594-3996
|
| Organization name |
NIH
|
| Department |
NCI-CCR
|
| Lab |
Laboratory of Cellular Oncology
|
| Street address |
37 Convent Dr.
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL15050 |
| Series (1) |
| GSE34770 |
ChIP-on-chip from murine myeloid cell line M1 and virus-induced myeloid leukemia cell lines for H3K4me3, H3K9/14ac, H3K4me1, H3K27me3, H3K9me3 and CTCF |
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