Sp Transcription Factor Western Blotting and ChIP

For Western blotting and ChIP of Sp1, Sp2 and Sp3 in MEFs, we used homemade rabbit antibodies [7 (link),42 (link)] affinity-purified against the respective recombinant Sp factor. Anti-Sp3 antibodies (Santa Cruz, sc-644) were used for the Western blot shown in Fig. 3B, and anti-Sp2 antibodies (Santa Cruz, sc-643) for ChIP-seq of Sp2 in HEK293 cells. Additional antibodies: Anti-Nf-ya (Santa Cruz, sc-10779), anti-Nf-yb (Genespin, PAb001), anti-Nf-yc (Santa Cruz, sc-7715-R), anti-Flag M2 (Sigma, F3165), anti-Tubulin (Millipore, MAB3408).

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Völkel S., Stielow B., Finkernagel F., Stiewe T., Nist A, & Suske G. (2015). Zinc Finger Independent Genome-Wide Binding of Sp2 Potentiates Recruitment of Histone-Fold Protein Nf-y Distinguishing It from Sp1 and Sp3. PLoS Genetics, 11(3), e1005102.

Publication 2015

Anti-Nf-ya anti-Flag M2 anti-Tubulin

Anti antibodies Antibodies Chip Chip seq Hek293 cells Rabbit Tubulin Western blot

Corresponding Organization :

Other organizations : Philipps University of Marburg

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Variable analysis

independent variables

Antibodies used for Western blotting and ChIP of Sp1, Sp2, and Sp3 in MEFs

dependent variables

Sp1, Sp2, and Sp3 protein levels (Western blotting)
Sp2 binding (ChIP-seq)

control variables

Cell type (MEFs)
Antibodies used for additional proteins (Nf-ya, Nf-yb, Nf-yc, Flag M2, Tubulin)

controls

Positive control: Recombinant Sp factors used for affinity purification of homemade antibodies
Negative control: Not explicitly mentioned

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