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. 2005 Mar 1;33(4):1269–1279. doi: 10.1093/nar/gki274

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© The Author 2005. Published by Oxford University Press. All rights reserved

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Immunodepletion of NELF or DSIF desensitizes transcription in vitro to DRB. (A) Mock-depleted (lanes 1 and 2) or NELF-depleted (lanes 3 and 4) extracts were tested for transcription in vitro in the presence or absence of DRB. NELF was depleted with a mixture of antibodies against NELF-D and NELF-E. Transcription was measured using an hsp70-G-less cassette substrate. The radiolabeled G-less transcription products are marked by a vertical line. The identity of the band marked with an asterisk is not known, but this band is evident even when transcription reactions are done in the presence of alpha amanitin (data not shown). It serves as a convenient control for RNA recovery. (B) Immunoblot analysis of proteins in mock-depleted and NELF-depleted extracts. (C) Mock-depleted (lanes 1 and 2) or DSIF-depleted (lanes 3 and 4) extracts were tested for transcription in vitro in the presence or absence of DRB. Antibody against the largest subunit of DSIF was used to deplete DSIF. (D) Immunoblot analysis of proteins in mock-depleted and DSIF-depleted extracts.