ChIP-PCR of CREB and SOX10

Cells were incubated with 1% formaldehyde (Sigma-Aldrich, F8775) to cross-link between DNA and proteins, and sonicated to yield chromatin fragments with about 200-500 base pairs. Chromatin fragments were reacted with anti-CREB antibody (Cell Signaling, 9197) or anti-SOX10 antibody (Santa Cruz, sc-17342) at 4℃ overnight, and precipitated with protein A-sepharose bead-sheared salmon sperm slurry (Merck, 17-295) for 4 h. Input and precipitated DNAs were subjected to PCR encompassing CREB- or SOX10-responsive element at the MITF-M promoter. Nucleotide sequences of PCR primers are previously described 35 (link). PCR products were resolved on agarose gels by electrophoresis and stained with EcoDye.

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Yun C.Y., Roh E., Kim S.H., Han J., Lee J., Jung D.E., Kim G.H., Jung S.H., Cho W.J., Han S.B, & Kim Y. (2020). Stem Cell Factor-Inducible MITF-M Expression in Therapeutics for Acquired Skin Hyperpigmentation. Theranostics, 10(1), 340-352.

Publication 2020

anti-CREB antibody anti-SOX10 antibody

Agarose Anti antibody Cells Chromatin Dnas Electrophoresis Formaldehyde Gels M element Mitf Nucleotide sequences Primers Protein a sepharose Proteins Salmon Sox10 Sperm

Corresponding Organization : Chungbuk National University

Other organizations : Chonnam National University, Chungnam National University

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Variable analysis

independent variables

Incubation with 1% formaldehyde (Sigma-Aldrich, F8775) to cross-link between DNA and proteins
Sonication to yield chromatin fragments with about 200-500 base pairs
Reaction with anti-CREB antibody (Cell Signaling, 9197) or anti-SOX10 antibody (Santa Cruz, sc-17342) at 4℃ overnight
Precipitation with protein A-sepharose bead-sheared salmon sperm slurry (Merck, 17-295) for 4 h

dependent variables

PCR encompassing CREB- or SOX10-responsive element at the MITF-M promoter
Resolution of PCR products on agarose gels by electrophoresis and staining with EcoDye

control variables

Incubation time (overnight)

controls

Positive control: Not specified
Negative control: Not specified

Annotations

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